Calcium channel drugs and uses

ABSTRACT

Novel multibinding compounds are disclosed. The compounds of this invention comprise 2-10 ligands covalently connected, each of the ligands being capable of binding to a ligand binding site in a Ca ++  channel , thereby modulating the biological activities thereof.

CONTINUING APPLICATION DATA

This on is a continuation-in-part of U.S. application Ser. No.09/325,557, filed Jun. 4, 1999, now abandoned, which, in turn, claimsthe benefit of U.S. Provisional Application Ser. No. 60/103,866 filedOct. 12, 1998.

BACKGROUND

1. Field of the Invention

This invention relates to novel multibinding compounds that bind to Ca⁺⁺channels and modulate their activity. The compounds of this inventioncomprise 2-10 Ca⁺⁺ channel ligands covalently connected by a linker orlinkers, wherein the ligands in their monovalent (i.e. unlinked) statebind to and are capable of modulating the activity of one or more typesof Ca⁺⁺ channel. The manner of linking the ligands together is such thatthe multibinding agents thus formed demonstrate an increased biologicand/or therapeutic effect as compared to the same number of unlinkedligands made available for binding to the Ca⁺⁺ channel. The inventionalso relates to methods of using such compounds and to methods ofpreparing them.

The compounds of this invention are particularly useful for treatingdiseases and conditions of mammals that are mediated by Ca⁺⁺ channels.Accordingly, this invention also relates to pharmaceutical compositionscomprising a pharmaceutically acceptable excipient and an effectiveamount of a compound of this invention.

2. State of the Art

Voltage-gated Ca⁺⁺ channels mediate the influx of Ca⁺⁺ into cells inresponse to changes in membrane potential. Because of their centralroles in ion homeostasis and in cell signaling events, these channelsare involved in a wide variety of physiological activities, e.g., musclecontraction, cardiovascular function, hormone and neurotransmittersecretion, and tissue growth and remodeling processes.

At least six types of calcium channels have been identified andcharacterized (Table 1, Appendix). The high-voltage activated Ca⁺⁺channels are formed by the heteromeric association of membrane proteinscomprising at least three subunits α (α₁, α₂), δ, β (and y in skeletalmuscle). The α₁ subunit alone is sufficient to form a functionalchannel, although the functional properties of the channel are subjectto modification, particularly by the β subunit. The α₁ subunit isorganized into four homologous domains (I-IV), each domain including 6transmembrane segments (S1-S6) (FIG. 1, Appendix). It is thought thatthe channel pore is formed from S5, S6 and the region between them, andthat the voltage sensor resides in S4.

These channels exist in resting (closed), activated (open) orinactivated (desensitized) states. The resting channels open in responseto depolarization of the membrane, then transition to an inactivatedstate.

Repolarization is required for return to the resting state. As shown inTable 1, channels differ in their activation and inactivationproperties.

Not surprisingly, Ca⁺⁺ channels are recognized as important targets fordrug therapy. They are implicated in a variety of pathologic conditions,including, e.g., essential hypertension, angina, congestive heartfailure, arrythmias, migraine and pain.

Calcium channel antagonists are potent vasodilators and are widely usedin the treatment of hypertension and angina pectoris. The compoundsapproved for clinical use in the U.S. fall into several chemicalclasses: the dihydropyridines (e.g., amlodipine, felodipine, nifedipine,nicardipine, isradipine, nimodipine); the benzothiazepines (e.g.,diltiazem), phenylalkylamines (e.g., verapamil); anddiarylaminopropylamine ether (e.g., bepridil).

The dihydropyridines, benzothiazepines and phenylalkylamines bind todistinct, but functionally coupled, sites on the α₁ subunit of L-typechannels at the interface of the IIIS6 and IVS6 transmembrane segments,such that the binding of any one class of drug can allostericallymodulate the binding of drugs in the other two classes and the highaffinity Ca⁺⁺ binding site in the channel (see G H Hockerman et al,Annu. Rev. Pharmacol. Toxicol. 37. 361-96 (1997)). It has been suggestedthat more than one high affinity binding site may exist fordihydropyridines in voltage-dependent calcium channels (Kokubun et al,Molec. Pharmacol. 30: 571-584 (1986)). However, studies reported in thescientific literature cast doubt on this hypothesis. In particular, theantagonist activities of a series of1,n-alkanediylbis(1,4-dihydropyridines) was reported to be essentiallyindependent of the bridging carbon chain length, and similar to that ofthe monomeric drugs (Joslyn et al., J. Med. Chem. 31: 1489-1492(1988)).

The clinical shortcomings of drugs in current usage are considerable.Various benzothiazepines and phenylalkylamines, for example, weakencardiac contractility and are therefore contraindicated in patients withleft ventricular dysfunction. Other Ca⁺⁺ channel antagonists cause AVblock, reflex tachycardia, excessive vasodilation and gastrointestinalproblems. Their most common adverse side effects include headache,flushing, hypotension, nausea, dizziness, fatigue, edema, abdominalpain, constipation, and the like. With few exceptions, the currentlyused drugs have a short duration of action and must be administeredfrequently for sustained effects.

Thus, there continues to exist a need for novel compounds with greatertissue selectivity, increased efficacy, reduced side effects and a morefavorable duration of action.

SUMMARY OF THE INVENTION

This invention is directed to novel multibinding compounds that bind toCa⁺⁺ channels in mammalian tissues and can be used to treat diseases andconditions mediated by such channels.

Accordingly, in one of its composition aspects, this invention isdirected to a multibinding compound and salts thereof comprising 2 to 10ligands which may be the same or different and which are covalentlyattached to a linker or linkers, which may be the same or different,each of said ligands comprising a ligand domain capable of binding to aCa⁺⁺ channel.

The multibinding compounds of this invention are preferably representedby Formula I:(L)_(p)(X)_(q)  I

-   where each L is a ligand that may be the same or different at each    occurrence;-   X is a linker that may be the same or different at each occurrence;-   p is an integer of from 2 to 10; and-   q is an integer of from 1 to 20;-   wherein each of said ligands comprises a ligand domain capable of    binding to a Ca⁺⁺ channel.    Preferably q is less than p.

More preferably the linker is represented by the following formula:—X′-Z-(Y′-Z)_(m)-Y″-Z-X′—in which:

-   m is an integer of from 0 to 20;-   X′ at each separate occurrence is —O—, —S—, —S(O)—, —S(O)₂—, —NR—,    —N⁺RR′—, —C(O)—, —C(O)O—, —C(O)NH—, —C(S), —C(S)O—, —C(S)NH— or a    covalent bond,-   where R and R′ at each separate occurrence are as defined below for    R′ and R″;

Z is at each separate occurrence selected from alkylene, substitutedalkylene, alkylalkoxy, cycloalkylene, substituted cycloalkylene,alkenylene, substituted alkenylene, alkynylene, substituted alkynylene,cycloalkenylene, substituted alkenylene, arylene, substituted arylene,heteroarylene, heterocyclene, substituted heterocyclene, crowncompounds, or a covalent bond;

Y′ and Y′ at each separate occurrence are selected from —S—S— or acovalent bond;

in which:

n is 0, 1 or 2; and

R′ and R″ at each separate occurrence are selected from hydrogen, alkyl,substituted alkyl, cycloalkyl, substituted cycloalkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, aryl, heteroaryl orheterocyclic.

Even more preferably, the ligands are selected from Ca⁺⁺ channelmodulators such as A-53930A, AE-0047, AGN-190604, AGN-190744, AH-1058,AHR-12742, AHR-16303B, AHR-16462B, AIT-110, AIT-111, AJ-3941, AM-336,amlopidine (including S(−), R-(+), and racemic), anipamil, AP-1067,aranidipine, atosiban, azelnidipine, barnidipine, Bay-t-7207,Bay-y-5959, Bay-z-4406, BBR-2160, belfosdil, BIII-890-CL, bisaramil,BMS-181102, BMS-188107, BMY-43011, BRL-32872, buflomedil, CD-349,CD-832, CERM-12816, CGP-28932, cilnidipine, dentiazem, clevidipine,CNS-1067, CNS-1237, CNS-2103, CP-060S, CPC-301, CPC-317, CPU-86017,D-2024, darodipine, DHP-218, diftiazem, diperdipine, dopropidil,dotarazine, dronedarone, DTZ-323, E-047/1, efonidipine, EGIS-7229,elgodipine, emopamil, etomoxir, F-0401, fantofarone, fasudil, FCE-24265,FCE-26262, FCE-27335, FCE-27892, FCE-28718, felodipine, FPL-64176,FR-172516, FRG-8701, fumidipine, GS-386, iganidipine, ipenoxazone,isradipine, JTV-591, KP-840, KT-362, L-366682, lacidipine, LAS-0538,LCB-2514, lemildipine, lercanidipine, leualacin, lifarizine, LOE-908,lomerizine, lubeluzole, LY-042826, manidipine, McN-6186, mibefradil,monatepil , MR-14134, N-3601, NCC-1048, nefiracetam, nexopamil,nifedipine, nifedipine, Nifelan, nilvadipine, nimodipine, NNC-09-0026,NPS-568, NS-638, NS-649, NS-696, NS-7, OPC-8490, Org-13061, Org-30029,oxodipine, P-5, palonidipine, PCA-50922, PCA-50938, PCA-50941,PD-029361, PD-157667, PD-158143, PD-176078, pranidipine, QX-314,ranolazine, RHG-2716, RingCap, Ro-11-2933, RS-5773, RU-43945, RWJ-22108,RWJ-22726, RWJ-29009, RWJ-37868, S-12968, S-2150, S-312-d, SANK-71996,SB-201823, SB-206284A, SB-23736, SD-3212, semotiadil, SIB-1281,siratiazem, SKFA-45675, SKF-96365, SKT-M-26, SL-34.0829, SL-87.0495,SM-6586, SNX-124, SNX-236, SNX-239, SNX-325, SNX-482, SQ-31727,SQ-33351, SQ-34399, SR-33805, TA-993, tamolarizine, TDN-345, temiverine,terodiline, TH-9229, TN-871, U-88999, U-92032, U-92798, UCL-1439,UK-1656, UK-55444, UK-56593, UK-84149, verapamil, Verelan, vexibinol,VUF-8929, WAY-141520, XB-513, XT-044, Y-22516, YH-334, YM-1615-4,YM-430, Z-6568, zatebradine, ziconotide, and ZM-224832.

Particularly preferred Ca⁺⁺ channel modulators include verapamil,diltiazem, benziazem clentiazem, nicardipine, nifedipine, nilvadipine,nitredipine, nimodipine, isradipine, lacidipine, amlodipine,nisoldipine, isradipine, mibefradil, amlodipine, felodipine, nimodipine,bepridil, SQ 32,910 and SQ 32,428.

In a second embodiment, this invention is directed to a pharmaceuticalcomposition comprising a pharmaceutically acceptable excipient and atherapeutically effective amount of one or more multibinding compounds(or pharmaceutically acceptable salts thereof) comprising 2 to 10ligands which may be the same or different and which are covalentlyattached to a linker or linkers, which may be the same or different,each of said ligands comprising a ligand domain capable of binding to aCa⁺⁺ channel of a cell mediating mammalian diseases or conditions,thereby modulating the diseases or conditions.

In a third embodiment, this invention is directed to a pharmaceuticalcomposition comprising a pharmaceutically acceptable excipient and atherapeutically effective amount of one or more multibinding compoundsrepresented by Formula I,(L)_(p)(X)_(q)  Ior pharmaceutically acceptable salts thereof,

-   where each L is a ligand that may be the same or different at each    occurrence;-   X is a linker that may be the same or different at each occurrence;-   p is an integer of from 2 to 10; and-   q is an integer of from 1 to 20;    wherein each of said ligands comprises a ligand domain capable of    binding to a Ca⁺⁺ channel of a cell mediating mammalian diseases or    conditions, thereby modulating the diseases or conditions,    Preferably q is less than p.

In a fourth embodiment, this invention is directed to a method formodulating the activity of a Ca⁺⁺ channel in a biologic tissue, whichmethod comprises contacting a tissue having a Ca⁺⁺ channel with amultibinding compound (or pharmaceutically acceptable salts thereof)under conditions sufficient to produce a change in the activity of thechannel in said tissue, wherein the multibinding compound comprises 2 to10 ligands which may be the same or different and which are covalentlyattached to a linker or linkers, which may be the same or different,each of said ligands comprising a ligand domain capable of binding to aCa⁺⁺ channel.

In a fifth embodiment, this invention is directed to a method fortreating a disease or condition in a mammal resulting from an activityof a Ca⁺⁺ channel, which method comprises administering to said mammal atherapeutically effective amount of a pharmaceutical compositioncomprising a pharmaceutically acceptable excipient and one or moremultibinding compounds (or pharmaceutically acceptable salts thereof)comprising 2 to 10 ligands which may be the same or different and whichare covalently attached to a linker or linkers, which may be the same ordifferent, each of said ligands comprising a ligand domain capable ofbinding to a Ca⁺⁺ channel of a cell mediating mammalian diseases orconditions.

In a sixth embodiment, this invention is directed to a method fortreating a disease or condition in a mammal resulting from an activityof a Ca⁺⁺ channel, which method comprises administering to said mammal atherapeutically effective amount of a pharmaceutical compositioncomprising a pharmaceutically acceptable excipient and one or moremultibinding compounds represented by Formula I,(L)_(p)(X)_(q)  Iand pharmaceutically acceptable salts thereof,

-   where each L is a ligand that may be the same or different at each    occurrence;-   X is a linker that may be the same or different at each occurrence;-   p is an integer of from 2 to 10; and-   q is an integer of from 1 to 20:    wherein each of said ligands comprises a ligand domain capable of    binding to a Ca⁺⁺ channel of a cell mediating mammalian diseases or    conditions. Preferably q is less than p.

In a seventh embodiment, this invention relates to processes forpreparing the multibinding agents of Formula I.

In an eighth aspect, this invention is directed to general syntheticmethods for generating large libraries of diverse multimeric compoundswhich multimeric compounds are candidates for possessing multibindingproperties. The diverse multimeric compound libraries provided by thisinvention are synthesized by combining a linker or linkers with a ligandor ligands to provide for a library of multimeric compounds wherein thelinker and ligand each have complementary functional groups permittingcovalent linkage. The library of linkers is preferably selected to havediverse properties such as valency, linker length, linker geometry andrigidity, hydrophilicity or hydrophobicity, amphiphilicity, acidity,basicity and polarization. The library of ligands is preferably selectedto have diverse attachment points on the same ligand, differentfunctional groups at the same site of otherwise the same ligand, and thelike.

This invention is also directed to libraries of diverse multimericcompounds which multimeric compounds are candidates for possessingmultibinding properties. These libraries are prepared via the methodsdescribed above and permit the rapid and efficient evaluation of whatmolecular constraints impart multibinding properties to a ligand or aclass of ligands targeting a receptor.

Accordingly, in one of its method aspects, this invention is directed toa method for identifying multimeric ligand compounds possessingmultibinding properties which method comprises:

(a) identifying a ligand or a mixture of ligands wherein each ligandcontains at least one reactive functionality;

(b) identifying a library of linkers wherein each linker in said librarycomprises at least two functional groups having complementary reactivityto at least one of the reactive functional groups of the ligand;

(c) preparing a multimeric ligand compound library by combining at leasttwo stoichiometric equivalents of the ligand or mixture of ligandsidentified in (a) wit the library of linkers identified in (b) underconditions wherein the complementary functional groups react to form acovalent linkage between said linker and at least two of said ligands;and

(d) assaying the multimeric ligand compounds produced in (c) above toidentify multimeric ligand compounds possessing multibinding properties.

In another of its method aspects, this invention is directed to a methodfor identifying multimeric ligand compounds possessing multibindingproperties which method comprises:

(a) identifying a library of ligands wherein each ligand contains atleast one reactive functionality;

(b) identifying a linker or mixture of linkers wherein each linkercomprises at least two functional groups having complementary reactivityto at least one of the reactive functional groups of the ligand;

(c) preparing a multimeric ligand compound library by combining at leasttwo stoichiometric equivalents of the library of ligands identified in(a) with the linker or mixture of linkers identified in (b) underconditions wherein the complementary functional groups react to form acovalent linkage between said linker and at least two of said ligands;and

(d) assaying the multimeric ligand compounds produced in (c) above toidentify multimeric ligand compounds possessing multibinding properties.

The preparation of the multimeric ligand compound library is achieved byeither the sequential or concurrent combination of the two or morestoichiometric equivalents of the ligands identified in (a) with thelinkers identified in (b). Sequential addition is preferred when amixture of different ligands is employed to ensure heterodimeric ormultimeric compounds are prepared. Concurrent addition of the ligandsoccurs when at least a portion of the multimer comounds prepared arehomomultimeric compounds.

The assay protocols recited in (d) can be conducted on the multimericligand compound library produced in (c) above, or preferably, eachmember of the library is isolated by preparative liquid chromatographymass spectrometry (LCMS).

In one of its composition aspects, this invention is directed to alibrary of multimeric ligand compounds which may possess multivalentproperties which library is prepared by the method comprising:

(a) identifying a ligand or a mixture of ligands wherein each ligandcontains at least one reactive functionality;

(b) identifying a library of linkers wherein each linker in said librarycomprises at least two functional groups having complementary reactivityto at least one of the reactive functional groups of the ligand; and

(c) preparing a multimeric ligand compound library by combining at leasttwo stoichiometric equivalents of the ligand or mixture of ligandsidentified in (a) with the library of linkers Identified in (b) underconditions wherein the complementary functional groups react to form acovalent linkage between said linker and at least two of said ligands.

In another of its composition aspects, this invention is directed to alibrary of multimeric ligand compounds which may possess multivalentproperties which library is prepared by the method comprising:

(a) identifying a library of ligands wherein each ligand contains atleast one reactive functionality;

(b) identifying a linker or mixture of linkers wherein each linkercomprises at least two functional groups having complementary reactivityto at least one of the reactive functional groups of the ligand; and

(c) preparing a multimeric ligand compound library by combining at leasttwo stoichiometric equivalents of the library of ligands identified in(a) with the linker or mixture of linkers identified in (b) underconditions wherein the complementary functional groups react to form acovalent linkage between said linker and at least two of said ligands.

In a preferred embodiment, the library of linkers employed in either themethods or the library aspects of this invention is selected from thegroup comprising flexible linkers, rigid linkers, hydrophobic linkers,hydrophilic linkers, linkers of different geometry, acidic linkers,basic linkers, linkers of different polarization and amphiphiliclinkers. For example, in one embodiment, each of the linkers in thelinker library may comprise linkers of different chain length and/orhaving different complementary reactive groups. Such linker lengths canpreferably range from about 2 to 100 Å.

In another preferred embodiment, the ligand or mixture of ligands isselected to have reactive functionality at different sites on saidligands in order to provide for a range of orientations of said ligandon said multimeric ligand compounds. Such reactive functionalityincludes, by way of example, carboxylic acids, carboxylic acid halides,carboxyl esters, amines, halides, isocyanates, vinyl unsaturation,ketones, aldehydes, thiols, alcohols, anhydrides, and precursorsthereof. It is understood, of course, that the reactive functionality onthe ligand is selected to be complementary to at least one of thereactive groups on the linker so that a covalent linkage can be formedbetween the linker and the ligand.

In other embodiments, the multimeric ligand compound is homomeric (ie.,each of the ligands is the same, although it may be attached atdifferent points) or heterodimeric (i.e., at least one of the ligands isdifferent from the other ligands).

In addition to the combinatorial methods described herein, thisinvention provides for an iterative process for rationally evaluatingwhat molecular constraints impart multibinding properties to a class ofmultimeric compounds or ligands targeting a receptor. Specifically, thismethod aspect is directed to a method for Identifying multimeric ligandcompounds possessing multibinding properties which method comprises:

(a) preparing a first collection or iteration of multimeric compoundswhich is prepared by contacting at least two stoichiometric equivalentsof the ligand or mixture of ligands which target a receptor with alinker or mixture of linkers wherein said ligand or mixture of ligandscomprises at least one reactive functionality and said linker or mixtureof linkers comprises at least two functional groups having complementaryreactivity to at least one of the reactive functional groups of theligand wherein said contacting is conducted under conditions wherein thecomplementary functional groups react to form a covalent linkage betweensaid linker and at least two of said ligands;

(b) assaying said first collection or iteration of multimeric compoundsto assess which if any of said multimeric compounds possess multibindingproperties;

(c) repeating the process of (a) and (b) above until at least onemultimeric compound is found to possess multibinding properties;

(d) evaluating what molecular constraints imparted multibindingproperties to the multimeric compound or compounds found in the firstiteration recited in (a)-(c) above;

(e) creating a second collection or iteration of multimeric compoundswhich elaborates upon the particular molecular constraints impartingmultibinding properties to the multimeric compound or compounds found insaid first iteration;

(f) evaluating what molecular constraints imparted enhanced multibindingproperties to the multimeric compound or compounds found in the secondcollection or iteration recited in (e) above;

(g) optionally repeating steps (e) and (f) to further elaborate uponsaid molecular constraints.

Preferably, steps (e) and (f) are repeated at least two times, morepreferably at from 2-50 times, even more preferably from 3 to 50 times,and still more preferably at least 5-50 times.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a highly schematic illustration of the transmembraneorganization of the α1 subunit of the voltage-gated Ca⁺⁺ channel.

FIG. 2 illustrates a method for optimizing the linker geometry forpresentation of ligands (filled circles) in bivalent compounds:

-   A. phenyldiacetylene core structure-   B. cyclohexane dicarboxylic acid core structure

FIG. 3 shows exemplary linker “core” structures.

FIGS. 4-20 illustrate convenient methods for preparing the multibindingcompounds of this invention.

FIG. 21 is a table of 741 calcium channel antagonists according to thepresent invention.

DETAILED DESCRIPTION OF THE INVENTION

Biological systems in general are controlled by molecular interactionsbetween bioactive ligands and their receptors, in which the receptor“reognizes” a molecule or a portion thereof (ie., a ligand domain) toproduce a biological effect. The voltage-gated Ca⁺⁺ channels areconsidered to be pharmacological receptors: they possess specificbinding sites for ligands having agonist and antagonist activities; thebinding of ligands to such sites allosterically modulates Ca⁺⁺ fluxthrough the channel; the channel properties (i.e., gating and ionselectivity) are regulatable; and various channels are known toassociate with G-proteins (D. Rampe and D. J. Triggle, Prog. Drug Res.40: 191-238 (1993). Accordingly, diseases or conditions that involve, orare mediated by, Ca⁺⁺ channels can be treated with pharmacologicallyactive ligands that interact with such channels to initiate, modulate orabrogate transporter activity.

The interaction of a Ca⁺⁺ channel and a Ca⁺⁺ channel-binding ligand maybe described in terms of “affinity” and “specificity”. The “affinity”and “specificity” of any given ligand-Ca⁺⁺ channel interaction isdependent upon the complementarity of molecular binding surfaces and theenergetic costs of complexation (i.e., the net difference in free energyΔG between bound and free states). Affinity may be quantified by theequilibrium constant of complex formation, the ratio of on/off rateconstants, and/or by the free energy of complex formation. Specificityrelates to the difference in binding affinity of a ligand for differentreceptors.

The net free energy of interaction of such ligands with a Ca⁺⁺ channelis the difference between energetic gains (enthalpy gained throughmolecular complementarity and entropy gained through the hydrophobiceffect) and energetic costs (enthalpy lost through decreased salvationand entropy lost through reduced translational, rotational andconformational degrees of freedom).

The compounds of this invention comprise 2 to 10 Ca⁺⁺ channel-bindingligands covalently linked together and capable of acting as multibindingagents. Without wishing to be bound by theory, the enhanced activity ofthese compounds is believed to arise at least in part from their abilityto bind in a multivalent manner with multiple ligand binding sites on aCa⁺⁺ channel or channels, which gives rise to a more favorable net freeenergy of binding. Multivalent interactions differ from collections ofindividual monovalent (univalent) interactions by being capable ofproviding enhanced biologic and/or therapeutic effect. Multivalentbinding can amplify binding affinities and differences in bindingaffinities, resulting in enhanced binding specificity as well asaffinity.

Definitions

As used herein:

The term “alkyl” refers to a monoradical branched or unbranchedsaturated hydrocarbon chain, preferably having from 1 to 40 carbonatoms, preferably 1-10 carbon atoms, more preferably 1-6 carbon atoms,such as methyl, ethyl, n-propyl, isopropyl, n-butyl, secondary butyl,tert-butyl, n-hexyl, n-octyl, n-decyl, n-dodecyl, 2-ethyldodecyl,tetradecyl, and the like, unless otherwise indicated.

The term “substituted alkyl” refers to an alkyl group as defined abovehaving from 1 to 5 substituents selected from the group consisting ofalkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino,aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl,keto, thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy,thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, aryl,aryloxy, heteroaryl, heteraryloxy, heterocyclic, heterocyclooxy,hydroxyamino, alkoxyamino, nitro, —SO-alkyl, —SO-aryl, —SO-heteroaryl,—SO₂-alkyl, —O₂-aryl, —SO₂-heteroaryl, and —NR^(a)R^(b), wherein R^(a)and R^(b) may be the same or different and and are chosen from hydrogen,optionally substituted alkyl, cycloalkyl, alkenyl, cycloalkenyl,alkynyl, aryl, heteroaryl and heterocyclic.

The term “alkylene” refers to a diradical of a branched or unbranchedsaturated hydrocarbon chain, preferably having from 1 to 40 carbonatoms, preferably 1-10 carbon atoms, more preferably 1-6 carbon atoms.This term is exemplified by groups such as methylene (—CH₂—), ethylene(—CH₂CH₂—), the propylene isomers (e.g., —CH₂CH₂CH₂— and —CH(CH₃)CH₂—)and the like.

The term “substituted alkylene” refers to an alkylene group as definedabove having from 1 to 5 substituents selected from the group consistingof alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, acyl, acylamino, acyloxy, amino,aminoacyl, aminoacyloxy, oxyacylamino, azido, cyano, halogen, hydroxyl,keto, thioketo, carboxyl, carboxylalkyl, thiol, thioalkoxy, substitutedthioalkoxy, aryl, aryloxy, thioaryloxy, heteroaryl, heteroaryloxy,thioheteroaryloxy, heterocyclic, heterocyclooxy, thioheterocyclooxy,nitro, and —NR^(a)R^(b), wherein R^(a) and R^(b) may be the same ordifferent and are chosen from hydrogen, optionally substituted alkyl,cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl andheterocydic. Additionally, such substituted alkylene groups includethose where 2 substituents on the alkylene group are fused to form oneor more cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, heterocyclic or heteroaryl groups fused to thealkylene group. The term “substituted alkylene” optionally includes analkylene chain as defined above in which the carbon chain is interruptedby one or more atoms chosen from O, S or N (e.g., ethers, sulfides andamines).

The term “alkaryl” or “aralkyl” refers to the groups -alkylene-aryl and-substituted alkylene-aryl in which alkylene and aryl are as definedherein. Such alkaryl groups are exemplified by benzyl, phenethyl and thelike.

The term “alkoxy” refers to the groups alkyl-O—, alkenyl-O—,cycloalkyl-O—, cycloalkenyl-O—, and alkynyl-O—, where alkyl, alkenyl,cycloalkyl, cycloalkenyl, and alkynyl are as defined herein. Preferredalkoxy groups are alkyl-O— and include, by way of example, methoxy,ethoxy, n-propoxy, iso-propoxy, n-butoxy, tert-butoxy, sec-butoxy,n-pentoxy, n-hexoxy, 1,2-dimethylbutoxy, and the like

The term “substituted alkoxy” refers to the groups substituted alkyl-O—,substituted alkenyl-O—, substituted cycloalkyl-O—, substitutedcycloalkenyl-O—, and substituted alkynyl-O— where substituted alkyl,substituted alkenyl, substituted cycloalkyl, substituted cycloalkenyland substituted alkynyl are as defined herein.

“Alkenyl” refers to a monoradical of a branched or unbranchedunsaturated hydrocarbon preferably having from 2 to 40 carbon atoms,preferably 2-10 carbon atoms, more preferably 2-6 carbon atoms, andpreferably having 1-6 double bonds. This term is further exemplified bysuch radicals as vinyl, prop-2-enyl, pent-3-enyl, hex-5-enyl,5-ethyldodec-3,6-dienyl, and the like.

The term “substituted alkenyl” refers to an alkenyl group as definedabove having from 1 to 5 substituents selected from the group consistingof alkoxy, substituted alkoxy, acyl, acylamino, acyloxy, amino,aminoacyl, aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl,keto, thioketo, carboxyl, carboxylalkyl, thiol, thioalkoxy, substitutedthioalkoxy, aryl, heteroaryl, heterocydic, aryloxy, thioaryloxy,heteroaryloxy, thioheteroaryloxy, heterocyclooxy, thioheterocyclooxy,nitro, —SO-alkyl, —SO-substituted alkyl, —SO-aryl, —SO-heteroaryl,—SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-aryl, —SO₂-heteroaryl, and—NR^(a)R^(b), wherein R^(a) and R^(b) may be the same or different andare chosen from hydrogen, optionally substituted alkyl, cycloalkyl,alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl and heterocyclic.

“Alkenylene” refers to a diradical of an unsaturated hydrocarbon,preferably having from 2 to 40 carbon atoms, preferably 2-10 carbonatoms, more preferably 2-6 carbon atoms, and preferably having 1-6double bonds. This term is further exemplified by such radicals as1,2-ethenyl, 1,3-prop-2-enyl, 1,5-pent-3enyl, 1,4-hex-5-enyl,5-ethyl-1,12-dodec-3,6-dienyl, and the like.

The term “substituted alkenylene” refers to an alkenylene group asdefined above having from 1 to 5 substituents, selected from the groupconsisting of alkoxy, substituted alkoxy, acyl, acylamino, acyloxy,amino, aminoacyl, aminoacyloxy, oxyacylamino, azido, cyano, halogen,hydroxyl, keto, thioketo, carboxyl, carboxylalkyl, thiol, thioalkoxy,substituted thioalkoxy, aryl, aryloxy, thioaryloxy, heteroaryl,heteroaryloxy, thioheteroaryloxy, heterocyclic, heterocyclooxy,thioheterocyclooxy, nitro, and NR^(a)R^(b), wherein R^(a) and R^(b) maybe the same or different and are chosen from hydrogen, optionallysubstituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl,heteroaryl and heterocyclic. Additionally, such substituted alkenylenegroups include those where 2 substituents on the alkenylene group arefused to form one or more cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, aryl, heterocyclic or heteroarylgroups fused to the alkenylene group.

“Alkynyl” refers to a monoradical of an unsaturated hydrocarbon,preferably having from 2 to 40 carbon atoms, preferably 2-10 carbonatoms, more preferably 2-6 carbon atoms, and preferably having 1-6triple bonds. This term is further exemplified by such radicals asacetylenyl, prop-2-ynyl, pent-3-ynyl, hex-5-ynyl,“ethyldodec-3,6-diynyl, and the like.

The term “substituted alkynyl” refers to an alkynyl group as definedabove having from 1 to 5 substituents, selected from the groupconsisting of alkoxy, substituted alkoxy, acyl, acylamino, acyloxy,amino, aminoacyl, aminoacyloxy, oxyacylamino, azido, cyano, halogen,hydroxyl, keto, thioketo, carboxyl, carboxylalkyl, thiol, thioalkoxy,substituted thioalkoxy, aryl, aryloxy, thioaryloxy, heteroaryl,heteroaryloxy, thioheteroaryloxy, heterocyclic, heterocyclooxy,thioheterocycloxy, nitro, —SO-alkyl, —SO-substituted alkyl, —SO-aryl,—SO-heteroaryl, —SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-aryl,—SO₂-heteroaryl, SO₂-heterocyclic, NR^(a)R^(b), wherein R^(a) and R^(b)may be the same or different and are chosen from hydrogen, optionallysubstituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl,heteroaryl and heterocyclic.

“Alkynylene” refers to a diradical of an unsaturated hydrocarbonradical, preferably having from 2 to 40 carbon atoms, preferably 2-10carbon atoms, more preferably 2-6 carbon atoms, and preferably having1-6 triple bonds. This term is further exemplified by such radicals as1,3-prop-2-ynyl, 1,5-pent-3-ynyl, 1,4-hex-5-ynyl,5-ethyl-1,12-dodec-3,6-diynyl, and the like.

The term “acyl” refers to the groups —CHO, alkyl-C(O)—, substitutedalkyl-C(O)—, cycloalkyl-C(O)—, substituted cycloalkyl-C(O)—,cycloalkenyl-C(O)—, substituted cycloalkenyl-C(O)—, aryl-C(O)—,heteroaryl-C(O)— and heterocyclic-C(O)— where alkyl, substituted alkyl,cycloalkyl, substituted cycloalkyl, cycloalkenyl, substitutedcycloalkenyl, aryl, heteroaryl and heterocyclic are as defined herein.

The term “acylamino” refers to the group —C(O)NRR where each R isindependently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl,heterocyclic or where both R groups are joined to form a heterocyclicgroup (e.g., morpholine) wherein alkyl, substituted alkyl, aryl,heteroaryl and heterocyclic are as defined herein.

The term “aminoacyl” refers to the group —NRC(O)R where each R isindependently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl, orheterocyclic wherein alkyl, substituted alkyl, aryl, heteroaryl andheterocyclic are as defined herein.

The term “aminoacyloxy” refers to the group —NRC(O)OR where each R isindependently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl, orheterocyclic wherein alkyl, substituted alkyl, aryl, heteroaryl andheterocyclic are as defined herein.

The term “acyloxy” refers to the groups alkyl-C(O)O—, substitutedalkyl-C(O)O—, cycloalkyl-C(O)O—, substituted cycloalkyl-C(O)O—,aryl-C(O)O—, heteroaryl-C(O)O—, and heterocyclic-C(O)O— wherein alkyl,substituted alkyl, cycloalkyl, substituted cycloalkyl, aryl, heteroaryl,and heterocyclic are as defined herein.

The term “aryl” refers to an unsaturated aromatic carbocydic group offrom 6 to 20 carbon atoms having a single ring (e.g., phenyl) ormultiple condensed (fused) rings (e.g., naphthyl or anthryl).

Unless otherwise constrained by the definition for the aryl substituent,such aryl groups can optionally be substituted with from 1 to 5substituents selected from the group consisting of acyloxy, hydroxy,thiol, acyl, alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl, cycloalkenyl,substituted alkyl, substituted alkoxy, substituted alkenyl, substitutedalkynyl, substituted cycloalkyl, substituted cycloalkenyl, amino,aminoacyl, acylamino, alkaryl, aryl, aryloxy, azido, carboxyl,carboxylalkyl, cyano, halo, nitro, heteroaryl, heteroaryloxy,heterocyclic, heterocyclooxy, aminoacyloxy, oxyacylamino, thioalkoxy,substituted thioalkoxy, thioaryloxy, thioheteroaryloxy, —SO-alkyl,—SO-substituted alkyl, —SO-aryl, —SO-heteroaryl, —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-aryl, —SO₂-heteroaryl, trihalomethyl,NR^(a)R^(b), wherein R^(a) and R^(b) may be the same or different andare chosen from hydrogen, optionally substituted alkyl, cycloalkyl,alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl and heterocyclic.Preferred aryl substituents include alkyl, alkoxy, halo, cyano, nitro,trihalomethyl, and thioalkoxy.

The term “aryloxy” refers to the group aryl-O— wherein the aryl group isas defined above including optionally substituted aryl groups as alsodefined above.

The term “arylene” refers to a diradical derived from aryl orsubstituted aryl as defined above, and is exemplified by 1,2-phenylene,1,3-phenylene, 1,4-phenylene, 1,2-naphthylene and the like.

The term “carboxyalkyl” refers to the group “—C(O)Oalkyl” where alkyl isas defined above.

The term “cycloalkyl” refers to cyclic alkyl groups of from 3 to 20carbon atoms having a single cyclic ring or multiple condensed rings.Such cycloalkyl groups include, by way of example, single ringstructures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclooctyl, andthe like, or multiple ring structures such as adamantanyl, and the like.

The term “substituted cycloalkyl” refers to cycloalkyl groups havingfrom 1 to 5 substituents selected from the group consisting of alkoxy,substituted alkoxy, cycloalkyl, cycloalkenyl, substituted cycloalkenyl,acyl, acylamino, acyloxy, amino, aminoacyl, aminoacyloxy, oxyaminoacyl,azido, cyano, halogen, hydroxyl, keto, thioketo, carboxyl,carboxylalkyl, thioaryloxy, thioheteroaryloxy, thioheterocyclooxy,thiol, thioalkoxy, substituted thioalkoxy, aryl, aryloxy, heteroaryl,heteroaryloxy, heterocyclic, heterocyclooxy, hydroxyamino, alkoxyamino,nitro, —SO-alkyl, —SO-substituted alkyl, —SO-aryl, —SO-heteroaryl,—SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-aryl, —SO₂-heteroaryl, andNR^(a)R^(b), wherein R^(a) and R^(b) may be the same or different andare chosen from hydrogen, optionally substituted alkyl, cycloalkyl,alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl and heterocyclic.

The term “cycloalkenyl” refers to cyclic alkenyl groups of from 4 to 20carbon atoms having a single cyclic ring or fused rings and at least onepoint of internal unsaturation. Examples of suitable cycloalkenyl groupsinclude, for instance, cyclobut-2-enyl, cyclopent-3-enyl,cyclooct-3-enyl and the like.

The term “substituted cycloalkenyl” refers to cycloalkenyl groups havingfrom 1 to 5 substituents selected from the group consisting of alkoxy,substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloalkenyl,substituted cycloalkenyl, acyl, acylamino, acyloxy, amino, aminoacyl,aminoacyloxy, oxyaminoacyl, azido, cyano, halogen, hydroxyl, keto,thioketo, carboxyl, carboxylalkyl, thioaryloxy, thioheteroaryloxy,thioheterocyclooxy, thiol, thioalkoxy, substituted thioalkoxy, arylaryloxy, heteroaryl, heteroaryloxy, heterocyclic, heterocyclooxy,hydroxyamino, alkoxyamino, nitro, —SO-alkyl, —SO-substituted alkyl,—SO-aryl, —SO-heteroaryl, —SO₂-alkyl, —SO₂-substituted alkyl, —SO₂-aryl,—SO₂-heteroaryl, and NR^(a)R^(b), wherein R^(a) and R^(b) may be thesame or different and are chosen from hydrogen, optionally substitutedalkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl, heteroaryl andheterocyclic.

The term “halo” or “halogen” refers to fluoro, chloro, bromo and iodo.

The term “heteroaryl” refers to an aromatic group of from 1 to 15 carbonatoms and 1 to 4 heteroatoms selected from oxygen, nitrogen and sulfurwithin at least one ring (if there is more than one ring).

Unless otherwise constrained by the definition for the heteroarylsubstituent, such heteroaryl groups can be optionally substituted with 1to 5 substituents selected from the group consisting of acyloxy,hydroxy, thiol, acyl, alkyl, alkoxy, alkenyl, alkynyl, cycloalkyl,cycloalkenyl, substituted alkyl, substituted alkoxy, substitutedalkenyl, substituted alkynyl, substituted cycloalkyl, substitutedcycloalkenyl, amino, aminoacyl, acylamino, alkaryl, aryl, aryloxy,azido, carboxyl, carboxylalkyl, cyano, halo, nitro, heteroaryl,heteroaryloxy, heterocyclic, heterocyclooxy, aminoacyloxy, oxyacylamino,thioalkoxy, substituted thioalkoxy, thioaryloxy, thioheteroaryloxy,—SO-alkyl, —SO-substituted alkyl, —SO-aryl, —SO-heteroaryl, —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-aryl, —SO₂-heteroaryl, trihalomethyl,mono-and di-alkylamino, mono- and NR^(a)R^(b), wherein R^(a) and R^(b)may be the same or different and are chosen from hydrogen, optionallysubstituted alkyl, cycloalkyl, alkenyl, cycloalkenyl, alkynyl, aryl,heteroaryl and heterocydic. Preferred heteroaryls nclude pyridyl,pyrrolyl and furyl.

The term “heteroaryloxy” refers to the group heteroaryl-O—.

The term “heteroarylene” refers to the diradical group derived fromheteroaryl or substituted heteroaryl as defined above, and isexemplified by the groups 2,6-pyridylene, 2,4-pyridiylene,1,2-quinolinylene, 1,8-quinolinylene, 1,4-benzofuranylene,2,5-pyridinylene, 1,3-morpholinylene, 2,5-indolenyl, and the like.

The term “heterocycle” or “heterocyclic” refers to a monoradicalsaturated or unsaturated group having a single ring or multiplecondensed rings, from 1 to 40 carbon atoms and from 1 to 10 heteroatoms, preferably 1 to 4 heteroatoms, selected from nitrogen, sulfur,phosphorus, and/or oxygen within the ring.

Unless otherwise constrained by the definition for the heterocyclicsubstituent, such heterocyclic groups can be optionally substituted with1 to 5, and preferably 1 to 3 substituents, selected from the groupconsisting of alkoxy, substituted alkoxy, cycloalkyl, substitutedcycloalkyl, cycloalkenyl, substituted cycloalkenyl, acyl, acylamino,acyloxy, amino, aminoacyl, aminoacyloxy, oxyaminoacyl, cyano, halogen,hydroxyl, keto, thioketo, carboxyl, carboxylalkyl, thioaryloxy,thioheteroaryloxy, thioheterocyclooxy, thiol, thioalkoxy, substitutedthioalkoxy, aryl, aryloxy, heteroaryl, heteroaryloxy, heterocyclic,heterocyclooxy, hydroxyamino, alkoxyamino, nitro, —SO-alkyl,—SO-substituted alkyl, —SO-aryl, —SO-heteroaryl, —SO₂-alkyl,—SO₂-substituted alkyl, —SO₂-aryl, —SO₂-heteroaryl, and NR^(a)R^(b),wherein R^(a) and R^(b) may be the same or different and are chosen fromhydrogen, optionally substituted alkyl, cycloalkyl, alkenyl,cycloalkenyl, alkynyl, aryl, heteroaryl and heterocyclic. Suchheterocyclic groups can have a single ring or multiple condensed rings.

Examples of nitrogen heterocycles and heteroaryls include, but are notlimited to, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrmidine,pyridazine, indolizine, isoindole, indole, indazole, purine,quinolizine, isoquinoline, quinoline, phthalazine, naphthylpyridine,quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline,phenanthridine, acridine, phenanthroline, isothiazole, phenazine,isoxazole, phenoxazine, phenothiazine, imidazolidine, imidazoline,piperidine, piperazine, indoline, morpholino, piperidinyl,tetrahydrofuranyl, and the like as well as N-alkoxy-nitrogen containingheterocycles.

A preferred class of heterocyclics include “crown compounds” whichrefers to a specific class of heterocyclic compounds having one or morerepeating units of the formula [—(CH₂—)_(m)Y—] where m is equal to orgreater than 2, and Y at each separate occurrence can be O, N, S or P.Examples of crown compounds include, by way of example only,[—(CH₂)₃—NH—]₃, [—((CH₂)₂—O)₄—((CH₂)₂)—NH)₂] and the like. Typicallysuch crown compounds can have from 4 to 10 heteroatoms and 8 to 40carbon atoms.

The term “heterocyclooxy” refers to the group heterocyclic-O—.

The term “thioheterocyclooxy” refers to the group heterocydic-S—.

The term “heterocyclene” refers to the diradical group derived from aheterocycle as defined herein, and is exemplified by the groups2,6-morpholino, 2,5-morpholino and the like.

The term “oxyacylamino” refers to the group —OC(O)NRR where each R isindependently hydrogen, alkyl, substituted alkyl, aryl, heteroaryl, orheterocyclic wherein alkyl, substituted alkyl, aryl, heteroaryl andheterocydic are as defined herein.

The term “thiol” refers to the group —SH.

The term “thioalkoxy” refers to the group —S-alkyl.

The term “substituted thioalkoxy” refers to the group —S-substitutedalkyl.

The term “thioaryloxy” refers to the group aryl-S— wherein the arylgroup is as defined above including optionally substituted aryl groupsalso defined above.

The term “thioheteroaryloxy” refers to the group heteroaryl-S— whereinthe heteroaryl group is as defined above including optionallysubstituted aryl groups as also defined above.

As to any of the above groups which contain one or more substituents, itis understood, of course, that such groups do not contain anysubstitution or substitution patterns which are sterically impracticaland/or synthetically non-feasible. In addition, the compounds of thisinvention include all stereochemical isomers arising from thesubstitution of these compounds.

“Alkyl optionally interrupted by 1-5 atoms chosen from O, S, or N”refers to alkyl as defined above in which the carbon chain isinterrupted by O, S, or N, Within the scope are ethers, sulfides, andamines, for example 1-methoxydecyl, 1-pentyloxynonane,1-(2-isopropoxyethoxy)-4-methylnonane, 1-(2-ethoxyethoxy)dodecyl,2-(t-butoxy)heptyl, 1-pentylsulfanylnonane, nonylpentylamine, and thelike.

“Heteroarylalkyl” refers to heteroaryl as defined above linked to alkylas defined above, for example pyrid-2-ylmethyl, 8-quinolinylpropyl, andthe like.

“Optional” or “optionally” means that the subsequently described eventor circumstance may or may not occur, and that the description includesinstances where said event or circumstance occurs and instances in whichit does not. For example, optionally substituted alkyl means that alkylmay or may not be substituted by those groups enumerated in thedefinition of substituted alkyl.

The term “pharmaceutically acceptable salt” refers to salts which retainthe biological effectiveness and properties of the multibindingcompounds of this invention and which are not biologically or otherwiseundesirable. In many cases, the multibinding compounds of this inventionare capable of forming acid and/or base salts by virtue of the presenceof amino and/or carboxyl groups or groups similar thereto.

Pharmaceutically acceptable base addition salts can be prepared frominorganic and organic bases. Salts derived from inorganic bases, includeby way of example only, sodium, potassium, lithium, ammonium, calciumand magnesium salts. Salts derived from organic bases include, but arenot limited to, salts of primary, secondary and tertiary amines, such asalkyl amines, dialkyl amines, trialkyl amines, substituted alkyl amines,di(substituted alkyl) amines, tri(substituted alkyl) amines, alkenylamines, dialkenyl amines, trialkenyl amines, substituted alkenyl amines,di(substituted alkenyl) amines, tri(substituted alkenyl) amines,cycloalkyl amines, di(cycloalkyl) amines, tri(cycloalkyl) amines,substituted cycloalkyl amines, disubstituted cycloalkyl amine,trisubstituted cycloalkyl amines, cycloalkenyl amines, di(cycloalkenyl)amines, tri(cycloalkenyl) amines, substituted cycloalkenyl amines,disubstituted cycloalkenyl amine, trisubstituted cycloalkenyl amines,aryl amines, diaryl amines, triaryl amines, heteroaryl amines,diheteroaryl amines, triheteroaryl amines, heterocyclic amines,diheterocyclic amines, triheterocyclic amines, mixed di- and tri-amineswhere at least two of the substituents on the amine are different andare selected from the group consisting of alkyl, substituted alkyl,alkenyl, substituted alkenyl, cycloalkyl, substituted cycloalkyl,cycloalkenyl, substituted cycloalkenyl, aryl, heteroaryl, heterocyclic,and the like. Also included are amines where the two or threesubstituents, together with the amino nitrogen, form a heterocyclic orheteroaryl group.

Examples of suitable amines include, by way of example only,isopropylamine, trimethyl amine, diethyl amine, tri(isopropyl) amine,tri(n-propyl) amine, ethanolamine, 2-dimethylaminoethanol, tromethamine,lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline,betaine, ethylenediamine, glucosamine, N-alkylglucamines, theobromine,purines, piperazine, piperidine, morpholine, N-ethylpiperdine, and thelike. It should also be understood that other carboxylic acidderivatives would be useful in the practice of this invention, forexample, carboxylic acid amides, including carboxamides, lower alkylcarboxamides, dialkyl carboxamides, and the like.

Pharmaceutically acceptable acid addition salts may be prepared frominorganic and organic acids. Salts derived from inorganic acids includehydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid, and the like. Salts derived from organic acids includeacetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid,malic acid, malonic acid, succinic acid, maleic acid, fumaric acid,tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid,methanesulfonic acid, ethanesulfonic acid, p-toluene-sulfonic acid,salicylic acid, and the like.

The term “library” refers to at least 3, preferably from 10² to 10⁹ andmore preferably from 10² to 10⁴ multimeric compounds. Preferably, thesecompounds are prepared as a multiplicity of compounds in a singlesolution or reaction mixture which permits facile synthesis thereof. Inone embodiment, the library of multimeric compounds can be directlyassayed for multibinding properties. In another embodiment, each memberof the library of multimeric compounds is first isolated and,optionally, characterized. This member is then assayed for multibindingproperties.

The term “collection” refers to a set of multimeric compounds which areprepared either sequentially or concurrently (e.g., combinatorially).The collection comprises at least 2 members: preferably from 2 to 10⁹members and still more preferably from 10 to 10⁴ members.

The term “multimeric compound” refers to compounds comprising from 2 to10 ligands covalently connected through at least one linker whichcompounds may or may not possess multibinding properties (as definedherein).

The term “pseudohalide” refers to functional groups which react indisplacement reactions in a manner similar to a halogen. Such functionalgroups include, by way of example, mesyl, tosyl, azido and cyano groups.

The term “protecting group” or “blocking group” refers to any groupwhich when bound to one or more hydroxyl, thiol, amino or carboxylgroups of the compounds prevents reactions from occurring at thesegroups and which protecting group can be removed by conventionalchemical or enzymatic steps to reestablish the hydroxyl, thiol, amino orcarboxyl group. See, generally, T. W. Greene & P. G. M. Wuts “ProtectiveGroups in Organic Synthesis,” 2^(nd) Ed, 1991, John Wiley and Sons, N.Y.

The particular removable blocking group employed is not critical andpreferred removable hydroxyl blocking groups include conventionalsubstituents such as allyl, benzyl, acetyl, chloroacetyl, thiobenzyl,benzylidine, phenacyl, t-butyl-diphenylsilyl and any other group thatcan be introduced chemically onto a hydroxyl functionality and laterselectively removed either by chemical or enzymatic methods in mildconditions compatible with the nature of the product.

Preferred removable amino blocking groups include conventionalsubstituents such as t-butyoxycarbonyl (t-BOC), benzyloxycarbonyl (CBZ),fluorenylmethoxycarbonyl (FMOC), allyloxycarbonyl (ALOC) and the like,which can be removed by conventional conditions compatible with thenature of the product.

Preferred carboxyl protecting groups include esters such as methyl,ethyl, propyl, t-butyl etc. which can be removed by mild hydrolysisconditions compatible with the nature of the product.

As used herein, the terms “inert organic solvent” or “inert solvent”mean a solvent inert under the conditions of the reaction beingdescribed in conjunction therewith [including, for example, benzene,toluene, acetonitrile, tetrahydrofuran (“THF”), dimethylformamide(“DMF”), chloroform (“CHCl_(3”)), methylene chloride (or dichloromethaneor “CH₂Cl₂”), diethyl ether, ethyl acetate, acetone, methylethyl ketone,methanol, ethanol, propanol, isopropanol, tert-butanol, dioxane,pyridine, and the like]. Unless specified to the contrary, the solventsused in the reactions of the present invention are inert solvents.

The term “Ca⁺⁺ channel” refers to a structure comprised of integralmembrane proteins that functions to allow Ca⁺⁺ to equilibrate across amembrane according to its electrochemical gradient and at rates that arediffusion-limited. Examples of various types of Ca⁺⁺ channels are givenin Table 1 (Appendix).

“Ligand” as used herein denotes a compound that is a binding partner fora Ca⁺⁺ channel receptor, and is bound thereto, for example, bycomplementarity. The specific region or regions of the ligand moleculethat is recognized by the ligand binding site of a Ca⁺⁺ channel receptoris designated as the “ligand domain”. A ligand may be either capable ofbinding to a receptor by itself, or may require the presence of one ormore non-ligand components for binding (e.g. ions, a lipid molecule, asolvent molecule, and the like).

Ligands useful in this invention comprise Ca⁺⁺ channel modulators suchas verapamil (a phenylalkylamine), diltiazem (a benzothiazepine),nicardipine, nifedipine, isradipine, amlodipine, mibefradil, felodipineand nimodipine (dihydropyridines), and bepridil (adiarylaminopropylamine ether). See Tables 4 and 5, Appendix forstructures of various dihydropyridine, phenylalkylamine, andbenzothiazepine ligands.

While it is contemplated that many calcium channel ligands that arecurrently known can be used in the preparation of multibinding compoundsof this invention (see Table 2 (Appendix)), it should be understood thatportions of the ligand structure that are not essential for molecularrecognition and binding activity (i.e., that are not part of the liganddomain) may be varied substantially, replaced with unrelated structuresand, in some cases, omitted entirely without affecting the bindinginteraction. Accordingly, it should be understood that the term “ligand”is not intended to be limited to compounds known to be useful as Ca⁺⁺channel receptor-binding compounds (e.g., known drugs), in that ligandsthat exhibit marginal activity or lack useful activity as monomers canbe highly active as multibinding compounds, because of the biologicalbenefit conferred by multivalency. The primary requirement for a ligandas defined herein is that it has a ligand domain, as defined above,which is available for binding to a recognition site on a Ca⁺⁺ channel.

For purposes of the present invention, the term “ligand” or “ligands” isintended to include the racemic ligands as well as the individualstereoisomers of the ligands, including pure enantiomers and non-racemicmixtures thereof. A large number of known calcium channel ligands haveat least one chiral center and show stereoselective pharmacokinetics andpharmacologic activity (reviewed in Tokuma, Y and Noguchi, H., J.Chromatography A, 694: 181-193 (1995)). The scope of the invention asdescribed and claimed encompasses the racemic forms of the ligands aswell as the individual enantiomers and non-racemic mixtures thereof.

The term “ligand binding site” as used herein denotes a site on a Ca⁺⁺channel receptor that recognizes a ligand domain and provides a bindingpartner for the ligand. The ligand binding site may be defined bymonomeric or multimeric structures. This interaction may be capable ofproducing a unique biological effect, for example agonism, antagonism,modulation, or may maintain an ongoing biological event, and the like.

It should be recognized that the ligand binding sites of Ca⁺⁺ channelreceptors that participate in biological multivalent bindinginteractions are constrained to varying degrees by their intra- andintermolecular associations. For example, Ca⁺⁺ channel ligand bindingsites may be covalently joined in a single structure, nonchalantlyassociated in one or more multimeric structures, embedded in a membraneor biopolymer matrix, and so on, and therefore have less translationaland rotational freedom than if the same sites were present as monomersin solution.

The terms “agonism” and “antagonism” are well known in the art. As usedherein, the term “agonist” refers to a ligand that when bound to a Ca⁺⁺channel stimulates its activity. The term “antagonist” refers to aligand that when bound to a Ca⁺⁺ channel inhibits its activity. Channelblock or activation may result from allosteric effects of ligand bindingto the channel rather than occupancy of the channel pore. Theseallosteric effects may produce changes in protein conformation thataffect Ca⁺⁺ binding sites, gating mechanisms and/or the pore region(i.e., ion permeation).

A channel can exist in three modes: mode 0 (where the channel has zeroprobability of opening); mode 1 (where the channel opens frequently buttransiently) and mode 2, where the channel remains open for relativelylong periods of time (Hess et al, Nature 311: 538-544 (1984)). Theprobability that a channel will exist in one of these three stateschanges with voltage. A given ligand may have different bindingaffinities for different states, and thereby be capable of producingagonist or antagonist activity.

The term “modulatory effect” is intended to refer to the ability of aligand to change the activity of a Ca⁺⁺ channel through binding to thechannel.

“Multibinding agent” or “multibinding compound” refers herein to acompound that has from 2 to 10 Ca⁺⁺ channel ligands as defined herein(which may be the same or different) covalenty bound to one or morelinkers (which may be the same or different), and is capable ofmultivalency, as defined below.

A multibinding compound provides an improved biologic and/or therapeuticeffect compared to that of the same number of unlinked ligands availablefor binding to the ligand binding sites on a Ca⁺⁺ channel or channels.Examples of improved “biologic and/or therapeutic effect” includeincreased ligand-receptor binding interactions (e.g., increasedaffinity, increased ability to elicit a functional change in the target,improved kinetics), increased selectivity for the target, increasedpotency, increased efficacy, decreased toxicity, increased therapeuticindex, improved duration of action, improved bioavailability, improvedpharmacokinetics, improved activity spectrum, and the like. Themultibinding compounds of this invention will exhibit at least one, andpreferably more than one, of the above-mentioned effects.

“Univalency” as used herein refers to a single binding interactionbetween one ligand with one ligand binding site as defined herein. Itshould be noted that a compound having multiple copies of a ligand (orligands) exhibits univalency when only one ligand of that compoundinteracts with a ligand binding site. Examples of univalent interactionsare depicted below.

“Multivalency” as used herein refers to the concurrent binding of from 2to 10 linked ligands, which may be the same or different, and two ormore corresponding ligand binding sites, which may be the same ordifferent. An example of trivalent binding is depicted below forillustrative purposes.

It should be understood that not all compounds that contain multiplecopies of a ligand attached to a linker necessarily exhibit thephenomena of multivalency, i.e., that the biologic and/or therapeuticeffect of the multibinding agent is greater than that of the same numberof unlinked ligands made available for binding to the ligand bindingsites. For multivalency to occur, the ligand domains of the ligands thatare linked together must be presented to their cognate ligand bindingsites by the linker or linkers in a specific manner in order to bringabout the desired ligand-orienting result, and thus produce amultibinding interaction.

The term “linker”, identified where appropriate by the symbol X, refersto a group or groups that covalently links from 2 to 10 ligands (asdefined above) in a manner that provides a multibinding compound capableof multivalency. The linker is a ligand-orienting entity that permitsattachment of multiple copies of a ligand (which may be the same ordifferent) thereto.

The term “linker” includes everything that is not considered to be partof the ligand, e.g., ancillary groups such as solubilizing groups,lipophilic groups, groups that alter pharmacodynamics orpharmacokinetics, groups that modify the diffusability of themultibinding compound, spacers that attach the ligand to the linker,groups that aid the ligandorienting function of the linker, for example,by imparting flexibility or rigidity to the linker as a whole, or to aportion thereof, and so on. The term “linker” does not, however, coversolid inert supports such as beads, glass particles, rods, and the like,but it is to be understood that the multibinding compounds of thisinvention can be attached to a solid support if desired, for example,for use in separation and purification processes and for similarapplications.

The extent to which the previously discussed enhanced activity ofmultibinding compounds is realized in this invention depends upon theefficiency with which the linker or linkers that joins the ligandspresents them to their array of ligand binding sites. Beyond presentingthese ligands for multivalent interactions with ligand binding sites,the linker spatially constrains these interactions to occur withindimensions defined by the linker.

The linkers used in this invention are selected to allow multivalentbinding of ligands to any desired ligand binding sites of a Ca⁺⁺channel, whether such sites are located interiorly (e.g., within achannel/translocation pore), both interiorly and on the periphery of achannel, at the boundary region between the lipid bilayer and thechannel, or at any intermediate position thereof. The preferred linkerlength will vary depending on the distance between adjacent ligandbinding sites, and the geometry, flexibility and composition of thelinker. The length of the linker will preferably be in the range ofabout 2 Å to about 100 Å, more preferably from about 2 Å to about 50 Åand even more preferably from about 3 Å to about 10 Å.

The ligands are covalently attached to the linker or linkers usingconventional chemical techniques. The reaction chemistries resulting insuch linkage are well known in the art and involve the use of reactivefunctional groups present on the linker and ligand. Preferably, thereactive functional groups on the linker are selected relative to thefunctional groups available on the ligand for coupling, or which can beintroduced onto the ligand for this purpose. Again, such reactivefunctional groups are well known in the art. For example, reactionbetween a carboxylic acid of either the linker or the ligand and aprimary or secondary amine of the ligand or the linker in the presenceof suitable well-known activating agents results in formation of anamide bond covalently linking the ligand to the linker; reaction betweenan amine group of either the linker or the ligand and a sulfonyl halideof the ligand or the linker results in formation of a sulfonamide bondcovalently linking the ligand to the linker; and reaction between analcohol or phenol group of either the linker or the ligand and an alkylor aryl halide of the ligand or the linker results in formation of anether bond covalenty linking the ligand to the linker. Table 3(Appendix) illustrates numerous reactive functional groups and theresulting bonds formed by reaction therebetween. Where functional groupsare lacking, they can be created by suitable chemistries that aredescribed in standard organic chemistry texts such as J. March,“Advanced Organic Chemistry”, 4^(th) Edition, (Wiley-interscience (NewYork), 1992.

The relative orientation in which the ligand domains are displayeddepends both on the particular point or points of attachment of theligands to the linker, and on the framework geometry. The determinationof where acceptable substitutions can be made on a ligand is typicallybased on prior knowledge of structure-activity relationships of theligand and/or congeners and/or structural information aboutligand-receptor complexes (e.g., X-ray crystallography. NMR, and thelike). Such positions and synthetic protocols for linkage are well knownin the art and can be determined by those with ordinary skill in the art(see Methods of Preparation and FIGS. 4-20 of the Appendix), Followingattachment of a ligand to the linker or linkers, or to a significantportion thereof (e.g., 2-10 atoms of linker), the linker-ligandconjugate may be tested for retention of activity in a relevant assaysystem (see Utility and Testing below for representative assays).

At present, it is preferred that the multibinding compound is a bivalentcompound in which two ligands are covalently linked, or a trivalentcompound, in which three ligands are covalently linked. Linker design isfurther discussed under Methods of Preparation.

“Potency” as used herein refers to the minimum concentration at which aligand is able to achieve a desirable biological or therapeutic effect.The potency of a ligand is typically proportional to its affinity forits receptor. In some cases, the potency may be non-linearly correlatedwith its affinity. In comparing the potency of two drugs, e.g., amultibinding agent and the aggregate of its unlinked ligand, thedose-response curve of each is determined under identical testconditions (e.g. in an in vitro or in vivo assay, in an appropriateanimal model). The finding that the multibinding agent produces anequivalent biologic or therapeutic effect at a lower concentration thanthe aggregate unlinked ligand (e.g. on a per weight, per mole or perligand basis) is indicative of enhanced potency.

“Selectivity” or “specificity” is a measure of the binding preferencesof a ligand for different receptors. The selectivity of a ligand withrespect to its target receptor relative to another receptor is given bythe ratio of the respective values of K_(d) (i.e., the dissociationconstants for each ligand-receptor complex) or, in cases where abiological effect is observed below the K_(d), the ratio of therespective EC₅₀s or IC₅₀s (i.e., the concentrations that produce 50% ofthe maximum response for the ligand interacting with the two distinctreceptors).

The term “treatment” refers to any treatment of a disease or conditionin a mammal, particularly a human, and includes:

(i) preventing the disease or condition from occurring in a subjectwhich may be predisposed to the condition but has not yet been diagnosedwith the condition and, accordingly, the treatment constitutesprophylactic treatment for the pathologic condition;

(ii) inhibiting the disease or condition, i.e., arresting itsdevelopment;

(iii) relieving the disease or condition, i.e., causing regression ofthe disease or condition; or

(iv) relieving the symptoms resulting from the disease or conditionwithout addressing the underlying disease or condition, e.g., relievingsymptoms of angina pectoris and other conditions of ischemia but not anunderlying cause such as, for example, atherosclerotic disease orhypertension.

The phrase “disease or condition which is modulated by treatment with amultibinding Ca⁺⁺ channel ligand” covers all disease states and/orconditions that are generally acknowledged in the art to be usefullytreated with a ligand for a Ca⁺⁺ channel in general, and those diseasestates and/or conditions that have been found to be usefully treated bya specific multibinding compound of our invention, i.e., the compoundsof Formula I. Such disease states include, by way of example only,hypertension, angina pectoris (particularly vasospastic angina andunstable angina), cerebral ischemia, cardiac arrythmias (particularly,arrythmias resulting from calcium-related changes in membrane potentialand conduction), cardiac hypertrophy due to systolic or diastolicoverload, congestive heart failure, migraine, Raynaud's disease, acuterenal failure due to prolonged renal ischemia, and the like.

The term “therapeutically effective amount” refers to that amount ofmultibinding compound that is sufficient to effect treatment, as definedabove, when administered to a mammal in need of such treatment. Thetherapeutically effective amount will vary depending upon the subjectand disease condition being treated, the weight and age of the subject,the severity of the disease condition, the manner of administration andthe like, which can readily be determined by one of ordinary skill inthe art.

The term “pharmaceutically acceptable excipient” is intended to includevehicles and carriers capable of being coadministered with amultibinding compound to facilitate the performance of its intendedfunction. The use of such media for pharmaceutically active substancesis well known in the art. Examples of such vehicles and carriers includesolutions, solvents, dispersion media, delay agents, emulsions and thelike. Any other conventional carrier suitable for use with themultibinding compounds also falls within the scope of the presentinvention.

Methods of Preparation

Linkers

The linker or linkers, when covalently attached to multiple copies ofthe ligands, provides a biocompatible, substantially non-immunogenicmultibinding compound. The biological activity of the multibinding Ca⁺⁺channel compound is highly sensitive to the geometry, composition, size,length, flexibility or rigidity, the presence or absence of anionic orcationic charge, the relative hydrophobicity/hydrophilicity, and similarproperties of the linker. Accordingly, the linker is preferably chosento maximize the biological activity of the compound. The linker may bebiologically “neutral,” i.e., not itself contribute any additionalbiological activity to the multibinding compound, or it may be chosen tofurther enhance the biological activity of the compound. In general, thelinker may be chosen from any organic molecule construct that orientstwo or more ligands for binding to the receptors to permit multivalency.In this regard, the linker can be considered as a “framework” on whichthe ligands are arranged in order to bring about the desiredligand-orienting result, and thus produce a multibinding compound.

For example, different orientations of ligands can be achieved byvarying the geometry of the framework (linker) by use of mono- orpolycyclic groups, such as aryl and/or heteroaryl groups, or structuresincorporating one or more carbon-carbon multiple bonds (alkenyl,alkenylene, alkynyl or alkynylene groups). The optimal geometry andcomposition of frameworks (linkers) used in the multibinding compoundsof this invention are based upon the properties of their intendedreceptors. For example, it is preferred to use rigid cyclic groups(e.g., aryl, heteroaryl), or non-rigid cyclic groups (e.g., cycloalkylor crown groups) to reduce conformational entropy when such may benecessary to achieve energetically coupled binding.

Different hydrophobic/hydrophilic characteristics of the linker as wellas the presence or absence of charged moieties can readily be controlledby the skilled artisan. For example, the hydrophobic nature of a linkerderived from hexamethylene diamine (H₂N(CH₂)₆NH₂) or related polyaminescan be modified to be substantially more hydrophilic by replacing thealkylene group with a poly(oxyalkylene) group such as found in thecommercially available “Jeffamines” (class of surfactants).

Different frameworks can be designed to provide preferred orientationsof the ligands. The identification of an appropriate framework geometryfor ligand domain presentation is an important first step in theconstruction of a multi binding agent with enhanced activity. Systematicspatial searching strategies can be used to aid in the identification ofpreferred frameworks through an iterative process. FIG. 2 (Appendix)illustrates a useful strategy for determining an optimal frameworkdisplay orientation for ligand domains and can be used for preparing thebivalent compounds of this invention. Various alternative strategiesknown to those skilled in the art of molecular design can be substitutedfor the one described here.

As shown in FIG. 2, the ligands (shown as filled circles) are attachedto a central core structure such as phenyldiacetylene (Panel A) orcyclohexane dicarboxylic acid (Panel B). The ligands are spaced apartfrom the core by an attaching moiety of variable lengths m and n. If theligand possesses multiple attachment sites (see discussion below), theorientation of the ligand on the attaching moiety may be varied as well.The positions of the display vectors around the central core structuresare varied, thereby generating a collection of compounds. Assay of eachof the individual compounds of a collection generated as described willlead to a subset of compounds with the desired enhanced activities(e.g., potency, selectivity). The analysis of this subset using atechnique such as Ensemble Molecular Dynamics will suggest a frameworkorientation that favors the properties desired.

The process may require the use of multiple copies of the same centralcore stricture or combinations of different types of display cores. Itis to be noted that core structures other than those shown here can beused for determining the optimal framework display orientation of theligands. The above-described technique can be extended to trivalentcompounds and compounds of higher-order valency.

A wide variety of linkers is commercially available (see, e.g., ChemSources USA and Chem Sources international; the ACD electronic database;and Chemical Abstracts). Many of the linkers that are suitable for usein this invention fall into this category. Others can be readilysynthesized by methods known in the art, and as described below.Examples of linkers include aliphatic moieties, aromatic moieties,steroidal moieties, peptides, and the like. Specific examples arepeptides or polyamides, hydrocarbons, aromatics, heterocyclics, ethers,lipids, cationic or anionic groups, or a combination thereof.

Examples are given below and in FIG. 3 (Appendix), but it should beunderstood that various changes may be made and equivalents may besubstituted without departing from the true spirit and scope of theinvention. For example, properties of the linker can be modified by theaddition or insertion of ancillary groups into the linker, for example,to change the solubility of the multibinding compound (in water, fats,lipids, biological fluids, etc.), hydrophobicity, hydrophilicity, linkerflexibility, antigenicity, stability, and the like. For example, theintroduction of one or more poly(ethylene glycol) (PEG) groups onto thelinker enhances the hydrophilicity and water solubility of themultibinding compound, increases both molecular weight and molecularsize and, depending on the nature of the unPEGylated linker, mayincrease the in vivo retention time. Further, PEG may decreaseantigenicity and potentially enhances the overall rigidity of thelinker.

Ancillary groups that enhance the water solubility/hydrophilicity of thelinker, and accordingly, the resulting multibinding compounds, areuseful in practicing this invention. Thus, it is within the scope of thepresent invention to use ancillary groups such as, for example, smallrepeating units of ethylene glycols, alcohols, polyols, (e.g., glycerin,glycerol propoxylate, saccharides, including mono-, oligosaccharides,etc.) carboxylates (e.g., small repeating units of glutamic acid,acrylic acid, etc.), amines (e.g., tetraethylenepentamine), and the liketo enhance the water solubility and/or hydrophilicity of themultibinding compounds of this invention. In preferred embodiments, theancillary group used to improve water solubility/hydrophilicity will bea polyether. In particularly preferred embodiments, the ancillary groupwill contain a small number of repeating ethylene oxide (—CH₂CH₂O—)units.

The incorporation of lipophilic ancillary groups within the structure ofthe linker to enhance the lipophilicity and/or hydrophobicity of thecompounds of Formula I is also within the scope of this invention.Lipophilic groups useful with the linkers of this invention include, butare not limited to, lower alkyl, aromatic groups and polycyclic aromaticgroups. The aromatic groups may be either unsubstituted or substitutedwith other groups, but are at least substituted with a group whichallows their covalent attachment to the linker. As used herein the term“aromatic groups” incorporates both aromatic hydrocarbons andheterocydic aromatics. Other lipophilic groups useful with the linkersof this invention include fatty acid derivatives which may or may notform micelles in aqueous medium and other specific lipophilic groupswhich modulate interactions between the multibinding compound andbiological membranes.

Also within the scope of this invention is the use of ancillary groupswhich result in the compound of Formula I being incorporated into avesicle, such as a liposome, or a micelle. The term “lipid” refers toany fatty acid derivative that is capable of forming a bilayer ormicelle such that a hydrophobic portion of the lipid material orientstoward the bilayer while a hydrophilic portion orients toward theaqueous phase. Hydrophilic characteristics derive from the presence ofphosphate, carboxylic, sulfato, amino, sulfhydryl, nitro and other likegroups well known in the art. Hydrophobicity could be conferred by theinclusion of groups that include, but are not limited to, long chainsaturated and unsaturated aliphatic hydrocarbon groups of up to 20carbon atoms and such groups substituted by one or more aryl,heteroaryl, cycloalkyl, and/or heterocyclic group(s). Preferred lipidsare phosphoglycerides and sphingolipids, representative examples ofwhich include phosphatidylcholine, phosphatidylethanolamine,phosphatidylserine, phosphatidylinositol, phosphatidic acid,palmitoyleoyl phosphatidylcholine, lysophosphatidylcholine,lysophosphatidyl-ethanolamine, dipalmitoylphosphatidylcholine,dioleoylphosphatidylcholine, distearoyl-phosphafidylcholine anddilinoleoylphosphatidylcholine. Other compounds lacking phosphorus, suchas sphingolipid and glycosphingolipid families, are also within thegroup designated as lipid. Additionally, the amphipathic lipidsdescribed above may be mixed with other lipids including triglyceridesand sterols.

The flexibility of the linker can be manipulated by the inclusion ofancillary groups which are bulky and/or rigid. The presence of bulky orrigid groups can hinder free rotation about bonds in the linker, orbonds between he linker and the ancillary group(s), or bonds between thelinker and the functional groups. Rigid groups can include, for example,those groups whose conformational freedom is restrained by the presenceof rings and/or n-bonds, for example, aryl, heteroaryl and heterocyclicgroups. Other groups which can impart rigidity include polypeptidegroups such as oligo- or polyproline chains.

Rigidity can also be imparted electrostatically. Thus, if the ancillarygroups are either positively or negatively charged, the similarlycharged ancillary groups will force the linker into a configurationaffording the maximum distance between each of the like charges. Theenergetic cost of bringing the like-charged groups closer to each other,which is inversely related to the square of the distance between thegroups, will tend to hold the linker in a configuration that maintainsthe separation between the like-charged ancillary groups. Further,ancillary groups bearing opposite charges will tend to be attracted totheir oppositely charged counterparts and potentially may enter intoboth inter- and intramolecular ionic bonds. This non-covalent mechanismwill tend to hold the linker in a conformation which allows bondingbetween the oppositely charged groups. The addition of ancillary groupswhich are charged, or alternatively, protected groups that bear a latentcharge which is unmasked, following addition to the linker, bydeprotection, a change in pH, oxidation, reduction or other mechanismsknown to those skilled in the art, is within the scope of thisinvention. Bulky groups can include, for example, large atoms, ions(e.g., iodine, sulfur, metal ions, etc.) or groups containing largeatoms, polycyclic groups, including aromatic groups, non-aromatic groupsand structures incorporating one or more carbon-carbon n-bonds (i.e.,alkenes and alkynes). Bulky groups can also include oligomers andpolymers which are branched- or straight-chain species. Species that arebranched are expected to increase the rigidity of the structure more perunit molecular weight gain than are straight-chain species.

In preferred embodiments, rigidity (entropic control) is imparted by thepresence of alicyclic (e.g., cycloalkyl), aromatic and heterocyclicgroups. In other preferred embodiments, this comprises one or moresix-membered rings. In still further preferred embodiments, the ring isan aryl group such as, for example, phenyl or naphthyl, or a macrocyclicring such as, for example, a crown compound.

In view of the above, it is apparent that the appropriate selection of alinker group providing suitable orientation, entropy andphysico-chemical properties is well within the skill of the art.

Eliminating or reducing antigenicity of the multibinding compoundsdescribed herein is also within the scope of this invention. In certaincases, the antigenicity of a multibinding compound may be eliminated orreduced by use of groups such as, for example, poly(ethylene glycol).

Compounds of Formula I

As explained above, the multibinding compounds described herein comprise2-10 ligands attached covalently to a linker that links the ligands in amanner that allows their multivalent binding to ligand binding sites ofCa⁺⁺ channels. The linker spatially constrains these interactions tooccur within dimensions defined by the linker. This and other factorsincreases the biologic and/or therapeutic effect of the multibindingcompound as compared to the same number of ligands used in monobindingform. The compounds of this invention are preferably represented by theempirical formula (L)_(p)(X)_(q) where L, X, p and q are as definedabove. This is intended to include the several ways in which the ligandscan be linked together in order to achieve the objective ofmultivalency, and a more detailed explanation is provided below.

As noted previously, the linker may be considered as a framework towhich ligands are attached. Thus, it should be recognized that theligands can be attached at any suitable position on this framework, forexample, at the termini of a linear chain or at any intermediateposition thereof.

The simplest and most preferred multibinding compound is a bivalentcompound which can be represented as L-X-L, where L is a ligand and isthe same or different and X is the linker. A trivalent compound couldalso be represented in a linear fashion, i.e., as a sequence of repeatedunits L-X-L-X-L, in which L is a ligand and is the same or different ateach occurrence, as is X. However, a trivalent compound can alsocomprise three ligands attached to a central core, and thus berepresented as (L)₃X, where the linker X could include, for example, anaryl or cycloalkyl group. Tetravalent compounds can be represented in alinear array, L-X-L-X-L-X-L, or a branched array,

i.e., a branched construct analogous to the isomers of butane (n-butyl,iso-butyl, sec-butyl, and t-butyl). Alternatively, it could berepresented as an aryl or cycloalkyl derivative as described above withfour (4) ligands attached to the core linker.

The same considerations apply to higher multibinding compounds of thisinvention containing from 5-10 ligands. However, for multibinding agentsattached to a central linker such as an aryl , cycloalkyl orheterocyclyl group, or a crown compound, there is a self-evidentconstraint that there must be sufficient attachment sites on the linkerto accommodate the number of ligands present; for example, a benzenering could not accommodate more than 6 ligands, whereas a multi-ringlinker (e.g., biphenyl) could accommodate a larger number of ligands.

The formula (L)_(p)(X)_(q) is also intended to represent a cycliccompound of formula (-L-X—)_(n), where n is 2-10.

All of the above variations are intended to be within the scope of theinvention defined by the formula (L)_(p)(X)_(q). Examples of bivalentand higher-order valency compounds of this invention are provided inFIGS. 4-20. With the foregoing in mind, a preferred linker may berepresented by the following formula:—X′-Z-(Y′-Z)_(m)-Y″-Z-X′—in which:

-   m is an integer of from 0 to 20;-   X′ at each separate occurrence is —O—, —S—, —S(O)—, —S(O)₂—, —NR—,    —N⁺RR′—, —C(O)—, —C(O)O—, —C(O)NH—, —C(S), —C(S)O—, —C(S)NH— or a    covalent bond, where R and R′ at each separate occurrence are as    defined below for R′ and R″,;

Z is at each separate occurrence selected from alkylene, substitutedalkylene, alkylalkoxy, cycloalkylene, substituted cycloalkylene,alkenylene, substituted alkenylene, alkynylene, substituted alkynylene,cycloalkenylene, substituted alkenylene, arylene, substituted arylene,heteroarylene, heterocyclene, substituted heterocyclene, crowncompounds, or a covalent bond;

Y′ and Y′ at each separate occurrence are selected from —S—S— or acovalent bond;

in which:

n is 0, 1 or 2; and

R′ and R″ at each separate occurrence are selected from hydrogen, alkyl,substituted alkyl, cycloalkyl, substituted cycloalkyl, alkenyl,substituted alkenyl, alkynyl, substituted alkynyl, aryl, heteroaryl orheterocyclic. Additionally, the linker moiety can be optionallysubstituted at any atom therein by one or more alkyl, substituted alkyl,cycloalkyl, substituted cycloalkyl, alkenyl, substituted alkenyl,alkynyl, substituted alkynyl, aryl, heteroaryl and heterocyclic group.

As indicated above, the simplest (and preferred) construct is a bivalentcompound which can be represented as L-X-L, where L is a Ca⁺⁺ channelligand that is the same or different at each occurrence, and X is thelinker. Accordingly, examples of the preparation of a bivalent ligandare given below as an illustration of the manner in which multibindingcompounds of Formula I are obtained.

The reaction schemes that follow illustrate preferred linking strategiesfor linking dihydropyridine, benzothiazepine and phenylalkylamineclasses of calcium channel modulators. These strategies are intended toapply as well to any Ca⁺⁺ channel ligand that includes, or can befunctionalized with groups compatible with the chosen linker (e.g.,mibefradil).

As was previously discussed, the linker or linkers can be attached todifferent positions on the ligand molecule to achieve differentorientations of the ligand domains and thereby facilitate multivalency.For example, the positions that are potentially available for linking adihydropyridine of formula (1), a benzothiazepine of formula (2)verapamil, a phenylalkylamine (3) and mibefradil, a tetralol derivativeand selective T-channel ligand (4) are indicated by arrows in thestructures shown below.

Preferred positions of attachment suggested by known SAR are illustratedin the reaction schemes of FIGS. 4-20. The R groups shown above arenumbered to correspond to structures shown in the reaction schemes.Examples of ligands are shown in Tables 4 and 5 (Appendix).

It should be understood that a large number of Ca⁺⁺ channel ligands arechiral and exhibit stereoselectivity. The most active enantiomers arepreferably used as ligands in the multibinding compounds of thisinvention. The chiral resolution of enantiomers is accomplished by wellknown procedures that result in the formation of diastereomericderivatives or salts, followed by conventional separation bychromatographic procedures or by fractional crystallization (see, e.g.,Bossert et al, Angew. Chem. Int. Ed. 20: 762-769 (1981) and U.S. Pat.No. 5,571,827 and references cited therein).

The ligands are covalently attached to the linker using conventionalchemical techniques. The reaction chemistries resulting in such linkageare well known in the art and involve the coupling of reactivefunctional groups present on the linker and ligand. In some cases, itmay be necessary to protect portions of the ligand that are not involvedin linking reactions. Protecting groups for this purpose are well knownin the art and are indicated generally in the reaction schemes by thesymbols PG and PG′. Preferably, the reactive functional groups on thelinker are selected relative to the functional groups on the ligand thatare available for coupling, or can be introduced onto the ligand forthis purpose. In some embodiments, the linker is coupled to ligandprecursors, with the completion of ligand synthesis being carried out ina subsequent step (see, e.g., FIG. 7, Appendix). Where functional groupsare lacking, they can be created by suitable chemistries that aredescribed in standard organic chemistry texts such as J. March,“Advanced Organic Chemistry”, 4^(th) Edition (Wiley-Interscience, N.Y.,1992). Examples of the chemistry for connecting ligands by a linker areshown in Table 3 (Appendix), where R¹ and R² represent a ligand and/orthe linking group. One skilled in the art will appreciate thatsynthetically equivalent coupling reactions can be substituted for thereactions illustrated herein.

The linker to which the ligands or ligand precursors are attachedcomprises a “core” molecule having two or more functional groups withreactivity that is complementary to that of the functional groups on theligand. FIG. 3 illustrates the diversity of “cores” that are useful forvarying the linker size, shape, length, orientation, rigidity,acidity/basicity, hydrophobicity/hydrophilicity, hydrogen bondingcharacteristics and number of ligands connected. This pictorialrepresentation is intended only to illustrate the invention, and not tolimit its scope to the structures shown. In the Figures and reactionschemes that follow, a solid circle is used to generically represent acore molecule. The solid circle is equivalent to a linker as definedabove after reaction.

Syntheses of Bivalent Compounds

The preferred compounds of Formula I are bivalent Accordingly, and forthe purpose of simplicity, the figures and reaction schemes belowillustrate the synthesis of bivalent Ca⁺⁺ channel modulators. It shouldbe noted, however, that the same techniques can be used to generatehigher order multibinding compounds, i.e., the compounds of theinvention where p is 3-10.

Reactions performed under standard amide coupling conditions are carriedout in an inert polar solvent (e.g., DMF, DMA) in the presence of ahindered base (e.g., TEA, DIPEA) and standard amide coupling reagents(e.g., DPPA, PyBOP, HATU, DCC).

Preparation of Dihydropyridine (DHP) Bivalent Compounds

FIG. 4 illustrates a preferred method for linking molecules at positionsR² (R⁶) of the dihydropyridine ring. As exemplified here for amlodipineand structurally analogous molecules, a multistep HantzschDHP-cyclization is used to introduce the appropriate groups [see, e.g.,Arrowsmith et al, J. Med. Chem. 29: 1696-1702 (1986); Bossert et al,Angew. Chem. Int. Ed. Engl. 20: 762-769 (1981)).

The starting materials are a compound of formula (4) having anazide-substituted β-keto ester and a substituted benzaldehyde (5). Thesecompounds are reacted in benzene in the presence of piperidine andacetic acid to yield an intermediate product, which, when reacted with acompound of formula (6) in benzene yields DHP (7), where R² isCH₂OCH₂CH₂N₃. The ring nitrogen is protected to yield the compound offormula (8), following which the azide is reduced to an amine compound(9) using hydrogen and an appropriate catalyst, such aspalladium/calcium carbonate. Amine (9) can then be coupled to a coremolecule using a variety of well known reactions, examples of which areshown in FIG. 4 and described below. The compound of formula (9) isreacted with a bifunctional alkylating core (e.g. a dibromide linker(10)) in an inert solvent (e.g. DMF) with a hindered base (e.g. DIPEA)to produce, after deprotection, the amine linked compound of Formula I(11). Compound (11) can also be formed by reacting (9) with a dialdehydecore (12) under standard reductive amination conditions (e.g. sodiumcyanoborohydride in ethanol with acid), followed by deprotection. In yetanother type of coupling reaction, the compound of form (9) is reactedwith an activated diacid core (13) in a polar inert solvent (e.g.CH₂Cl₂) to yield, after deprotection, the amide-linked compound ofFormula I (14). The diacid may be preactivated (e.g. by using the acidchloride), or activated in situ using conventional coupling conditions(e.g. DCC, DMAP, THF). Alternatively, the compound of formula (9) can bereacted with a diisocyanate core (15) in an inert solvent (e.g. THF) toyield, after deprotection, the urea-linked compound of Formula I (16).The compound of formula (9) can also be reacted with an activatedsulfonate core (17) in an inert solvent (e.g. THF) with a hindered base(e.g. DIPEA) to yield, after deprotection, the sulfonamide linkedcompound of Formula I (18).

FIGS. 5 and 6 illustrate the preparation of dihydropyridine compounds ofFormula I having different functional linking groups at positions R²(R⁶) of the dihydropyridine ring.

As shown in FIG. 5, a dihydropyridine with an alcohol side chain at R²is prepared by reacting a compound of formula (19) with (5) and (6)(using the same conditions as above), to make a compound of formula(20). After protection with a suitable amine-protecting group, thecompound of formula (21) can be coupled to various cores, as exemplifiedin FIG. 5. Alternatively, a compound of formula (21) can be prepared asdescribed in Alker, D and Denton, S. M., Tetrahedron, 46, 3693-3702,(1990).

The compound of formula (25) is reacted with a bifunctional alkylatingcore (e.g., a diol (10)) in an inert solvent (e.g. THF) with a strongdeprotonating base (e.g. NaH) to produce, after deprotection, an etherlinked compound of Formula I (22). Alternatively, the compound offormula (21) is reacted with an activated diacid core (13) in a polarinert solvent (e.g. THF) with base to yield, after deprotection, anester-linked compound of Formula I (23). Alternatively, the compound offormula (21) is reacted with a diisocyanate core (15) in an inertsolvent (e.g. THF) to yield, after deprotection, a carbamic ester-linkedcompound of Formula I (24).

FIG. 6 illustrates reactions for converting a nucleophilic side chaininto an electrophilic one. For example, the compound of formula (21) isreacted under standard chlorinating conditions (e.g. SO₂Cl₂ in thepresence of a suitable base such as imidazole in DMF) to yield chloride(25). The compound of formula (21) is reacted with mesyl chloride in aninert solvent (e.g., THF) in the presence of a base to yield mesylate(26). The compound of formula (21) is reacted with oxidating agents(e.g. CrO₃) to yield acid (27). The compound of form (21) is reactedunder mild oxidating conditions (e.g. PCC/CH₂Cl₂) to yield aldehyde(28).

The electrophilic groups thus generated can then be used in standardcoupling reactions such as those shown in FIGS. 4 and 5, for example,the reaction illustrated in FIG. 6 (bottom). In this reaction, a molarequivalent of a diol core (29) is reacted with 2 molar equivalents of amesylated dihydropyridine (26) in an inert solvent with a base to yield,after deprotection, an ether-linked compound of Formula I (30).

FIG. 7 illustrates a preferred method for forming bivalent DHPcompounds, which involves coupling of the linker to ligand precursors,followed by synthesis of the ligand. As shown here, a molar equivalentof a diol core (29) is coupled with approximately two molar equivalentsof a chloride-substituted β diketone (31) in an inert solvent (e.g. DMF)in the presence of a strong base (e.g. NaH) to yield an ether-linkedcompound of formula (32). Compound (32) is reacted with (5) and (6), asdescribed above with reference to FIGS. 4 and 5, to form an ether-linkedcompound of Formula I (30).

Dihydropyridine dimers may be linked through an ester linkage at R⁷ orR⁸, as is shown in FIG. 8. The starting material, a t-butyl ester offormula (33) is reacted with compounds (5) and (6) as describedpreviously with reference to FIGS. 4 and 5, to yield a dihydropyridineof formula (34). Cleavage of the t-butyl group of (34) with dilute acidyields (35), which is then amine-protected to form (36). Standardactivation techniques (e.g. DCC/DMAP/FHF) are used to couple the acid toa nucleophile core as previously described, for example to a diol core(29), to yield, after deprotection, an ester-linked compound of FormulaI (37).

FIG. 9 illustrates another procedure for synthesizing an ester-inkeddihydropyridine compound of Formula I. Here, a molar equivalent of adiol core (29) is coupled with two molar equivalents of a diketonecompound of formula (38) in an acid-catalyzed transesterificationreaction to form a compound of formula (39). Reaction of (39) with (5)and (6), as described above, yields ester-linked (37).

Preparation of Benzothiazepine (BZT) Bivalent Compounds

Several methods for preparing bivalent benzothiazepine compounds areillustrated in the reaction schemes shown in FIG. 10 (Appendix).According to FIG. 10, the starting material, a compound of formula (41)is prepared as described in U.S. Pat. No. 4,552,695. Compound (41) isreacted with a dihalide core (10). In an inert solvent (e.g. THF) in thepresence of a strong base (e.g. NaH) to form an amide N-linked compoundof Formula I (42). Following the linking step, the R¹² side chain can bedeprotected if necessary and acylated as described in theabove-mentioned patent.

Alternative coupling reactions are possible, including those shown inFIG. 4. In such instances, those skilled in the art will know how tomodify the reaction conditions to compensate for the reducednucleophilicity of the amide nitrogen.

Alternatively, the starting material is a compound of formula (43)(where R¹² is an ether-protected hydroxyl group), which is prepared asdescribed in the above-referenced patent. Deprotection of this compoundyields the alcohol (44), which can be reacted with an activated diacidcore (13) in an inert solvent (e.g. DMF) to yield ester-inked (45) asshown here. Alternative coupling reactions can be used, such as thoseshown in FIG. 5.

Alternatively, a compound of formula (77) may be prepared as describedin the above-referenced patent. Treatment of compound (77) with BBr₃ inCH₂Cl₂ affords cleavage of the aryl ether to phenol (78). Compound (78)can be selectively alkylated at the phenolic hydroxyl group by reactionwith dihalide (10) in K₂CO₃/acetone solution to yield ether-linked (79).

Preparation of Phenylalkylamine (PAA) Bivalent Compounds

FIG. 11 illustrates methods for linking PAA molecules (as exemplified byverapamil) (mibefradil is also considered herein to be part of thecategory of phenylalkylamines). Phenyl acetonitrile (80) is treated witha basic condensing agent (e.g. sodium amide) in an inert solvent (e.g.toluene). N-protected (81) is slowly added to the amidine salt, asdescribed in U.S. Pat. No. 3,261,859. After deprotection, compound (82)is coupled to an electrophilic core, for example, with a dihalide core(10) in an inert solvent (e.g. DMF) in the presence of a hindered base(e.g. DIPEA) to form an amine-linked compound of Formula I (83). Othercoupling reactions may be substituted for the one shown here, e.g.,those shown in FIG. 4.

Alternatively, the amidine salt of (80), is coupled to O-protected (84).After deprotection, the resulting compound (85) is reacted with adihalide core (10), in a K₂CO₃/acetone solution to form an ether linkedcompound of Formula I (86). Other coupling reactions such as those shownin FIG. 5 may be substituted for the one shown here.

The strategies for preparing compounds of Formula I discussed aboveinvolve coupling the ligand directly to a homobifunctional core. Anotherstrategy that can be used with all ligands, and for the preparation ofboth bivalent and higher order multibinding compounds, is to introduce a‘spacer’ before coupling to a central core. Such a spacer can itself beselected from the same set as the possible core compounds. Examples ofthis linking strategy are shown in FIGS. 12 and 13, where the spacer isrepresented by a gray circle. As defined herein, the linker comprisesthe spacer+core.

Referring to FIG. 12, a dihydropyridine compound of formula (21),synthesized according to FIG. 5 above, is coupled to aheterobifunctional spacer (46) having an electrophilic group (e.g., Br)and a masked nucleophilic group (e.g., protected alcohol). Theprotecting groups, PG and PG′ are different (e.g., PG is Boc and PG′ isCbz) and are capable of selective removal. The reaction is carried outin an inert solvent (e.g. DMF) in the presence of a strong base (e.g.NaH). After removal of PG′, the unmasked nucleophile (49) is coupled toan activated diacid core (13). The ring nitrogen is then deprotected toyield an ether-linked compound of Formula I (48). Core molecules withdifferent functional groups may be substituted for the one shown here(see, e.g., FIG. 5).

In another example, a dihydropyridine compound of formula (36) iscoupled to heterobifunctional spacer (49) under standard couplingconditions (e.g. DCC/DMAP/CH₂Cl₂). After removal of the spacerprotecting group, the unmasked nucleophile (50) is coupled to adialdehyde core (12) by reductive amination (or to another core withchemically compatible functional groups). After removal of theprotecting group from the ring nitrogen, an ester-inked compound offormula I (51) is obtained.

FIG. 13 Illustrates the use of spacers to make bivalent benzothiazepinecompounds. As shown in FIG. 13, a compound of formula (78) is coupled toa heterobifunctional spacer (46) in basic conditions (e.g. K₂CO₃ inacetone). After removal of the spacer protecting group, the unmaskednucleophile (52) is coupled to an activated diacid core (13) usingstandard coupling conditions to yield an ether-linked compound ofFormula I (53). In another example, compound (41) is coupled toheterobifunctional spacer (54) in an inert solvent solvent (e.g. DMF) inthe presence of strong base (e.g. NaH). After removal of the spacerprotecting group, the resulting compound (55) is coupled to an activateddiacid core (13) to yield an N-alkylated compound of Formula I (56). Ofcourse, other suitable cores may be substituted if desired.

Compounds of Formula I where p=3-10

Compounds of Formula I of higher order valency, i.e. p>2, can beprepared by simple extension of the above strategies. As shown in FIG.14 (Appendix), compounds (58) and (61) are prepared by coupling ligandsto a central core bearing multiple functional groups. The reactionconditions are the same as described above for the preparation ofbivalent compounds, with appropriate adjustments made in the molarquantities of ligand and reagents.

Compound (36), synthesized as described above with reference to FIG. 5.is coupled to a polypeptide core with a sidechain spacer (62) to make(63). Solid phase peptide synthesis can be used to produce a widevariety of peptidic core molecules. Techniques well known to thoseskilled in the art (including combinatorial methods) are used to varythe distance between ligand attachment sites on the core molecule, thenumber of attachment sites available for coupling, and the chemicalproperties of the core molecule. Orthogonal protecting groups are usedto selectively protect functional groups on the core molecule, thusallowing ancillary groups to be inserted into the linker of themultibinding compound and/or the preparation of “heterovalomers” (i.e.,multibinding compounds with nonidentical ligands). All of the syntheticstrategies described above employ a step in which the ligand, attachedto spacers or not, is symmetrically linked to functionally equivalentpositions on a central core. Compounds of Formula I can also besynthesized using an asymmetric linear approach. This strategy ispreferred when linking two or more ligands at different points ofconnectivity (see, e.g., FIG. 15) or when preparing heterovalomers (see,e.g., FIGS. 16 and 17).

FIG. 15 illustrates the preparation of bivalent dihydropyridinecompounds of formulae (64) and (65), wherein R² of a first ligand isattached through a linker to R⁷ of a second ligand, and the preparationof a bivalent benzothiazepine compound of formula (67), wherein thelinker is attached between R¹¹ of a first ligand and R¹³ of a secondligand. The coupling steps are carried out as described previously.

A linear strategy can also be used to prepare heterovalomers, as shownin FIGS. 16 and 17. Heterovalomers comprising different chemical classesof Ca⁺⁺ channel ligands (e.g., dihydropyridines and benzothiazepines),different ligands within the same chemical class (e.g. amlodipine andisradipine) and different enantiomers of a ligand (e.g., the (+) and (−)enantiomers of a dihydropyridine) are all encompassed by the presentinvention.

FIG. 16 illustrates methods of preparing bivalent compounds comprisingdihydropyridine and benzothiazepine ligands in which the orientation ofthe ligands is varied.

FIG. 17 illustrates the preparation of a mixed agonist/antagonistheterovalomer. In this example, the compound of formula (66), a Ca⁺⁺channel antagonist with an attached spacer, is coupled to the compoundof formula (75), a Ca⁺⁺ channel agonist, and deprotected to yield acompound of formula (76).

FIGS. 18-20 illustrate alternate methods of preparing bivalent compoundscomprising dihydropyridine and benzothiazepine derivatives.

Isolation and Purification of the Compounds

Isolation and purification of the compounds and intermediates describedherein can be effected, if desired, by any suitable separation orpurification such as, for example, filtration, extraction,crystallization, column chromatography, thin-layer chromatography,thick-layer chromatography, preparative low or high-pressure liquidchromatography or a combination of these procedures. Characterization ispreferably by NMR and mass spectroscopy.

Combinatorial Libraries

The methods described above lend themselves to combinatorial approachesfor identifying multimeric compounds which possess multibindingproperties.

Specifically, factors such as the proper juxtaposition of the individualligands of a multibinding compound with respect to the relevant array ofbinding sites on a target or targets is important in optimizing theinteraction of the multibinding compound with its target(s) and tomaximize the biological advantage through multivalency. One approach isto identify a library of candidate multibinding compounds withproperties spanning the multibinding parameters that are relevant for aparticular target. These parameters include: (1) the identity ofligand(s), (2) the orientation of ligands, (3) the valency of theconstruct, (4) linker length, (5) linker geometry, (6) linker physicalproperties, and (7) linker chemical functional groups. Libraries ofmultimeric compounds potentially possessing multibinding properties(i.e., candidate multibinding compounds) and comprising a multiplicityof such variables are prepared and these libraries are then evaluatedvia conventional assays corresponding to the ligand selected and themultibinding parameters desired. Considerations relevant to each ofthese variables are set forth below:

Selection of ligand(s):

A single ligand or set of ligands is (are) selected for incorporationinto the libraries of candidate multibinding compounds which library isdirected against a particular biological target or targets. The onlyrequirement for the ligands chosen is that they are capable ofinteracting with the selected target(s). Thus, ligands may be knowndrugs, modified forms of known drugs, substructures of known drugs orsubstrates of modified forms of known drugs (which are competent tointeract with the target), or other compounds. Ligands are preferablychosen based on known favorable properties that may be projected to becarried over to or amplified in multibinding forms. Favorable propertiesinclude demonstrated safety and efficacy in human patients, appropriatePK/ADME profiles, synthetic accessibility, and desirable physicalproperties such as solubility, log P. etc. However, it is crucial tonote that ligands which display an unfavorable property from among theprevious list may obtain a more favorable property through the processof multibinding compound formation; i.e., ligands should not necessarilybe excluded on such a basis. For example, a ligand that is notsufficiently potent at a particular target so as to be efficacious in ahuman patient may become highly potent and efficacious when presented inmultibinding form. A ligand that is potent and efficacious but not ofutility because of a non-mechanism-related toxic side effect may haveincreased therapeutic index (increased potency relative to toxicity) asa multibinding compound. Compounds that exhibit short in vivo half-livesmay have extended half-lives as multibinding compounds. Physicalproperties of ligands that limit their usefulness (e.g. poorbioavailability due to low solubility, hydrophobicity, hydrophilicity)may be rationally modulated in multibinding forms, providing compoundswith physical properties consistent with the desired utility.

Orientation: Selection of Ligand Attachment Points and Linking Chemistry

Several points are chosen on each ligand at which to attach the ligandto the linker. The selected points on the ligand/linker for attachmentare functionalized to contain complementary reactive functional groups.This permits probing the effects of presenting the ligands to theirreceptor(s) in multiple relative orientations, an important multibindingdesign parameter. The only requirement for choosing attachment points isthat attaching to at least one of these points does not abrogateactivity of the ligand. Such points for attachment can be identified bystructural information when available. For example, inspection of aco-crystal structure of a protease inhibitor bound to its target allowsone to identify one or more sites where linker attachment will notpreclude the enzyme:inhibitor interaction. Alternatively, evaluation ofligand/target binding by nuclear magnetic resonance will permit theidentification of sites non-essential for ligand/target binding. See,for example, Fesik, et al., U.S. Pat. No. 5,891,643. When suchstructural information is not available, utilization ofstructure-activity relationships (SAR) for ligands will suggestpositions where substantial structural variations are and are notallowed. In the absence of both structural and SAR information, alibrary is merely selected with multiple points of attachment to allowpresentation of the ligand in multiple distinct orientations. Subsequentevaluation of this library will indicate what positions are suitable forattachment.

It is important to emphasize that positions of attachment that doabrogate the activity of the monomeric ligand may also be advantageouslyincluded in candidate multibinding compounds in the library providedthat such compounds bear at least one ligand attached in a manner whichdoes not abrogate intrinsic activity. This selection derives from, forexample, heterobivalent interactions within the context of a singletarget molecule. For example, consider a receptor antagonist ligandbound to its target receptor, and then consider modifying this ligand byattaching to it a second copy of the same ligand with a linker whichallows the second ligand to interact with the same receptor molecule atsites proximal to the antagonist binding site, which include elements ofthe receptor that are not part of the formal antagonist binding siteand/or elements of the matrix surrounding the receptor such as themembrane. Here, the most favorable orientation for interaction of thesecond ligand molecule with the receptor/matrix may be achieved byattaching it to the linker at a position which abrogates activity of theligand at the formal antagonist binding site. Another way to considerthis is that the SAR of individual ligands within the context of amultibinding structure is often different from the SAR of those sameligands in momomeric form.

The foregoing discussion focused on bivalent interactions of dimericcompounds bearing two copies of the same ligand joined to a singlelinker through different attachment points, one of which may abrogatethe binding/activity of the monomeric ligand. It should also beunderstood that bivalent advantage may also be attained withheterodimeric constructs bearing two different ligands that bind tocommon or different targets. For example, a 5HT₄ receptor antagonist anda bladder-selective muscarinic M₃ antagonist may be joined to a linkerthrough attachment points which do not abrogate the binding affinity ofthe monomeric ligands for their respective receptor sites. The dimericcompound may achieve enhanced affinity for both receptors due tofavorable interactions between the 5HT₄ ligand and elements of the M₃receptor proximal to the formal M₃ antagonist binding site and betweenthe M₃ ligand and elements of the 5HT₄ receptor proximal to the formal5HT₄ antagonist binding site. Thus, the dimeric compound may be morepotent and selective antagonist of overactive bladder and a superiortherapy for urinary urge incontinence.

Once the ligand attachment points have been chosen, one identifies thetypes of chemical linkages that are possible at those points. The mostpreferred types of chemical linkages are those that are compatible withthe overall structure of the ligand (or protected forms of the ligand)readily and generally formed, stable and intrinsically inocuous undertypical chemical and physiological conditions, and compatible with alarge number of available linkers. Amide bonds, ethers, amines,carbamates, ureas, and sulfonamides are but a few examples of preferredlinkages.

Linkers: Spanning Relevant Multibinding Parameters through Selection ofValency, Linker Length, Linker Geometry, Rigidity, Physical Properties,and Chemical Functional Groups

In the library of linkers employed to generate the library of candidatemultibinding compounds, the selection of linkers employed in thislibrary of linkers takes into consideration the following factors:

Valency:

In most instances the library of linkers is initiated with divalentlinkers. The choice of ligands and proper juxtaposition of two ligandsrelative to their binding sites permits such molecules to exhibit targetbinding affinities and specificities more than sufficient to conferbiological advantage. Furthermore, divalent linkers or constructs arealso typically of modest size such that they retain the desirablebiodistribution properties of small molecules.

Linker Length:

Linkers are chosen in a range of lengths to allow the spanning of arange of inter-ligand distances that encompass the distance preferablefor a given divalent interaction. In some instances the preferreddistance can be estimated rather precisely from high-resolutionstructural information of targets, typically enzymes and solublereceptor targets. In other instances where high-resolution structuralinformation is not available (such as 7TM G-protein coupled receptors),one can make use of simple models to estimate the maximum distancebetween binding sites either on adjacent receptors or at differentlocations on the same receptor. In situations where two binding sitesare present on the same target (or target subunit for multisubunittargets), preferred linker distances are 2-20, with more preferredlinker distances of 3-12. In situations where two binding sites resideon separate (e.g., protein) target sites, preferred linker distances are20-100, with more preferred distances of 30-70

Linker Geometry and Rigidity:

The combination of ligand attachment site, linker length, linkergeometry, and linker rigidity determine the possible ways in which theligands of candidate multibinding compounds may be displayed in threedimensions and thereby presented to their binding sites. Linker geometryand rigidity are nominally determined by chemical composition andbonding pattern, which may be controlled and are systematically variedas another spanning function in a multibinding array. For example,linker geometry is varied by attaching two ligands to the ortho, meta,and para positions of a benzene ring, or in cis- or trans-arrangementsat the 1,1- vs. 1,2- vs. 1,3- vs. 1,4-positions around a cyclohexanecore or in cis- or trans-arrangements at a point of ethyleneunsaturation. Linker rigidity is varied by controlling the number andrelative energies of different conformational states possible for thelinker. For example, a divalent compound bearing two ligands joined by1,8-octyl linker has many more degrees of freedom, and is therefore lessrigid than a compound in which the two ligands are attached to the 4,4′positions of a biphenyl linker.

Linker Physical Properties:

The physical properties of linkers are nominally determined by thechemical constitution and bonding patterns of the linker, and linkerphysical properties impact the overall physical properties of thecandidate multibinding compounds in which they are included. A range oflinker compositions is typically selected to provide a range of physicalproperties (hydrophobicity, hydrophilicity, amphiphilicity,polarization, acidity, and basicity) in the candidate multibindingcompounds. The particular choice of linker physical properties is madewithin the context of the physical properties of the ligands they joinand preferably the goal is to generate molecules with favorable PK/ADMEproperties. For example, linkers can be selected to avoid those that aretoo hydrophilic or too hydrophobic to be readily absorbed and/ordistributed in vivo.

Linker Chemical Functional Groups:

Linker chemical functional groups are selected to be compatible with thechemistry chosen to connect linkers to the ligands and to impart therange of physical properties sufficient to span initial examination ofthis parameter.

Combinatorial Synthesis:

Having chosen a set of n ligands (n being determined by the sum of thenumber of different attachment points for each ligand chosen) and mlinkers by the process outlined above, a library of (n!)m candidatedivalent multibinding compounds is prepared which spans the relevantmultibinding design parameters for a particular target. For example, anarray generated from two ligands, one which has two attachment points(A1, A2) and one which has three attachment points (B1, B2, B3) joinedin all possible combinations provide for at least 15 possiblecombinations of multibinding compounds:

A1-A1 A1-A2 A1-B1 A1-B2 A1-B3 A2-A2 A2-B1 A2- B2 A2-B3 B1-B1 B1-B2 B1-B3B2-B2 B2-B3 B3-B3When each of these combinations is joined by 10 different linkers, alibrary of 150 candidate multibinding compounds results.Given the combinatorial nature of the library, common chemistries arepreferably used to join the reactive functionaries on the ligands withcomplementary reactive functionalities on the linkers. The librarytherefore lends itself to efficient parallel synthetic methods. Thecombinatorial library can employ solid phase chemistries well known inthe art wherein the ligand and/or linker is attached to a solid support.Alternatively and preferably, the combinatorial libary is prepared inthe solution phase. After synthesis, candidate multibinding compoundsare optionally purified before assaying for activity by, for example,chromatographic methods (e.g., HPLC).Analysis of Array by Biochemical, Analytical, Pharmacological, andComputational Methods:

Various methods are used to characterize the properties and activitiesof the candidate multibinding compounds in the library to determinewhich compounds possess multibinding properties. Physical constants suchas solubility under various solvent conditions and logD/clog D valuescan be determined. A combination of NMR spectroscopy and computationalmethods is used to determine low-energy conformations of the candidatemultibinding compounds in fluid media. The ability of the members of thelibrary to bind to the desired target and other targets is determined byvarious standard methods, which include radioligand displacement assaysfor receptor and ion channel targets, and kinetic inhibition analysisfor many enzyme targets. In vitro efficacy, such as for receptoragonists and antagonists, ion channel blockers, and antimicrobialactivity, can also be determined. Pharmacological data, including oralabsorption, everted gut penetration, other pharmacokinetic parametersand efficacy data can be determined in appropriate models. In this way,key structure-activity relationships are obtained for multibindingdesign parameters which are then used to direct future work.

The members of the library which exhibit multibinding properties, asdefined herein, can be readily determined by conventional methods. Firstthose members which exhibit multibinding properties are identified byconventional methods as described above including conventional assays(both in vitro and in vivo).

Second, ascertaining the structure of those compounds which exhibitmultibinding properties can be accomplished via art recognizedprocedures. For example, each member of the library can be encrypted ortagged with appropriate information allowing determination of thestructure of relevant members at a later time. See, for example, Dower,et al., International Patent Application Publication No. WO 93/06121;Brenner, et al., Proc. Natl. Acad. Sci., USA, 89:5181 (1992); Gallop, etal., U.S. Pat. No. 5,846,839; each of which are incorporated herein byreference in its entirety. Alternatively, the structure of relevantmultivalent compounds can also be determined from soluble and untaggedlibraries of candidate multivalent compounds by methods known in the artsuch as those described by Hindsgaul, et al., Canadian PatentApplication No. 2,240,325 which was published on Jul. 11, 1998. Suchmethods couple frontal affinity chromatography with mass spectroscopy todetermine both the structure and relative binding affinities ofcandidate multibinding compounds to receptors.

The process set forth above for dimeric candidate multibinding compoundscan, of course, be extended to trimeric candidate compounds and higheranalogs thereof.

Follow-Up Synthesis and Analysis of Additional Array(s):

Based on the information obtained through analysis of the initiallibrary, an optional component of the process is to ascertain one ormore promising multibinding “lead” compounds as defined by particularrelative ligand orientations, linker lengths, linker geometries, etc.Additional libraries can then be generated around these leads to providefor further information regarding structure to activity relationships.These arrays typically bear more focused variations in linker structurein an effort to further optimize target affinity and/or activity at thetarget (antagonism, partial agonism, etc.), and/or alter physicalproperties. By iterative redesign/analysis using the novel principles ofmultibinding design along with classical medicinal chemistry,biochemistry, and pharmacology approaches, one is able to prepare andidentify optimal multibinding compounds that exhibit biologicaladvantage towards their targets and as therapeutic agents.

To further elaborate upon this procedure, suitable divalent linkersinclude, by way of example only, those derived from dicarboxylic acids,disulfonylhalides, dialdehydes, diketones, dihalides, diisocyanates,diamines, diols, mixtures of carboxylic acids, sulfonylhalides,aldehydes, ketones, halides, isocyanates, amines and diols. In eachcase, the carboxylic acid, sulfonylhalide, aldehyde, ketone, halide,isocyanate, amine and diol functional group is reacted with acomplementary functionality on the ligand to form a covalent linkage.Such complementary functionality is well known in the art as illustratedin the following table:

COMPLEMENTARY BINDING CHEMISTRIES First Reactive Group Second ReactiveGroup Linkage hydroxyl isocyanate urethane amine epoxide -aminehydroxyamine sulfonyl halide sulfonamide carboxyl acid amine amidehydroxyl alkyl/aryl halide ether aldehyde amine/NaCNBH₄ amine ketoneamine/NaCNBH₄ amine amine isocyanate urea

The following table illustrates, by way of example, starting materials(identified as X-1 through X-418) that can be used to prepare linkersincorporated in the multibinding compounds of this invention utilizingthe chemistry described above. For example, 1,10-decanedicarboxylicacid, X1, can be reacted with 2 equivalents of a ligand carrying anamino group in the presence of a coupling reagent such as DCC to providea bivalent multibinding compound of formula (I) wherein the ligands arelinked via a 1,10-decanediamido linking group.

Representative ligands for use in this invention include, by way ofexample, L-1 through L-21, wherein L-1=verapamil, L-2=diltiazem,L-3=benziazem, L-4=clentiazem, L-5=nicardipine, L-6=nifedipine,L-7=nilvadipine, L-8=nitredipine, L-9=nimodipine, L-10=isradipine,L-11=lacidipine, L-12=amlodipine, L-13 nisoldipine, L-14=isradipine,L-15=mibefrodil, L-16=amlodipine, L-17=felodipine, L-18=nimodipine,L-19-bepridil, L-20=SQ32,910, and L-21-SQ32,248.

Combinations of ligands (L) and linkers (X) per this invention include,by way example only, homo- and hetero-dimers wherein a first ligand isselected from L-1 through L-21 above and the second ligand and linker isselected from the following:

L-1/X-1- L-1/X-2- L-1/X-3- L-1/X-4- L-1/X-5- L-1/X-6- L-1/X-7- L-1/X-8-L-1/X-9- L-1/X-10- L-1/X-11- L-1/X-12- L-1/X-13- L-1/X-14- L-1/X-15-L-1/X-16- L-1/X-17- L-1/X-18- L-1/X-19- L-1/X-20- L-1/X-21- L-1/X-22-L-1/X-23- L-1/X-24- L-1/X-25- L-1/X-26- L-1/X-27- L-1/X-28- L-1/X-29-L-1/X-30- L-1/X-31- L-1/X-32- L-1/X-33- L-1/X-34- L-1/X-35- L-1/X-36-L-1/X-37- L-1/X-38- L-1/X-39- L-1/X-40- L-1/X-41- L-1/X-42- L-1/X-43-L-1/X-44- L-1/X-45- L-1/X-46- L-1/X-47- L-1/X-48- L-1/X-49- L-1/X-50-L-1/X-51- L-1/X-52- L-1/X-53- L-1/X-54- L-1/X-55- L-1/X-56- L-1/X-57-L-1/X-58- L-1/X-59- L-1/X-60- L-1/X-61- L-1/X-62- L-1/X-63- L-1/X-64-L-1/X-65- L-1/X-66- L-1/X-67- L-1/X-68- L-1/X-69- L-1/X-70- L-1/X-71-L-1/X-72- L-1/X-73- L-1/X-74- L-1/X-75- L-1/X-76- L-1/X-77- L-1/X-78-L-1/X-79- L-1/X-80- L-1/X-81- L-1/X-82- L-1/X-83- L-1/X-84- L-1/X-85-L-1/X-86- L-1/X-87- L-1/X-88- L-1/X-89- L-1/X-90- L-1/X-91- L-1/X-92-L-1/X-93- L-1/X-94- L-1/X-95- L-1/X-96- L-1/X-97- L-1/X-98- L-1/X-99-L-1/X-100- L-1/X-101- L-1/X-102- L-1/X-103- L-1/X-104- L-1/X-105-L-1/X-106- L-1/X-107- L-1/X-108- L-1/X-109- L-1/X-110- L-1/X-111-L-1/X-112- L-1/X-113- L-1/X-114- L-1/X-115- L-1/X-116- L-1/X-117-L-1/X-118- L-1/X-119- L-1/X-120- L-1/X-121- L-1/X-122- L-1/X-123-L-1/X-124- L-1/X-125- L-1/X-126- L-1/X-127- L-1/X-128- L-1/X-129-L-1/X-130- L-1/X-131- L-1/X-132- L-1/X-133- L-1/X-134- L-1/X-135-L-1/X-136- L-1/X-137- L-1/X-138- L-1/X-139- L-1/X-140- L-1/X-141-L-1/X-142- L-1/X-143- L-1/X-144- L-1/X-145- L-1/X-146- L-1/X-147-L-1/X-148- L-1/X-149- L-1/X-150- L-1/X-151- L-1/X-152- L-1/X-153-L-1/X-154- L-1/X-155- L-1/X-156- L-1/X-157- L-1/X-158- L-1/X-159-L-1/X-160- L-1/X-161- L-1/X-162- L-1/X-163- L-1/X-164- L-1/X-165-L-1/X-166- L-1/X-167- L-1/X-168- L-1/X-169- L-1/X-170- L-1/X-171-L-1/X-172- L-1/X-173- L-1/X-174- L-1/X-175- L-1/X-176- L-1/X-177-L-1/X-178- L-1/X-179- L-1/X-180- L-1/X-181- L-1/X-182- L-1/X-183-L-1/X-184- L-1/X-185- L-1/X-186- L-1/X-187- L-1/X-188- L-1/X-189-L-1/X-190- L-1/X-191- L-1/X-192- L-1/X-193- L-1/X-194- L-1/X-195-L-1/X-196- L-1/X-197- L-1/X-198- L-1/X-199- L-1/X-200- L-1/X-201-L-1/X-202- L-1/X-203- L-1/X-204- L-1/X-205- L-1/X-206- L-1/X-207-L-1/X-208- L-1/X-209- L-1/X-210- L-1/X-211- L-1/X-212- L-1/X-213-L-1/X-214- L-1/X-215- L-1/X-216- L-1/X-217- L-1/X-218- L-1/X-219-L-1/X-220- L-1/X-221- L-1/X-222- L-1/X-223- L-1/X-224- L-1/X-225-L-1/X-226- L-1/X-227- L-1/X-228- L-1/X-229- L-1/X-230- L-1/X-231-L-1/X-232- L-1/X-233- L-1/X-234- L-1/X-235- L-1/X-236- L-1/X-237-L-1/X-238- L-1/X-239- L-1/X-240- L-1/X-241- L-1/X-242- L-1/X-243-L-1/X-244- L-1/X-245- L-1/X-246- L-1/X-247- L-1/X-248- L-1/X-249-L-1/X-250- L-1/X-251- L-1/X-252- L-1/X-253- L-1/X-254- L-1/X-255-L-1/X-256- L-1/X-257- L-1/X-258- L-1/X-259- L-1/X-260- L-1/X-261-L-1/X-262- L-1/X-263- L-1/X-264- L-1/X-265- L-1/X-266- L-1/X-267-L-1/X-268- L-1/X-269- L-1/X-270- L-1/X-271- L-1/X-272- L-1/X-273-L-1/X-274- L-1/X-275- L-1/X-276- L-1/X-277- L-1/X-278- L-1/X-279-L-1/X-280- L-1/X-281- L-1/X-282- L-1/X-283- L-1/X-284- L-1/X-285-L-1/X-286- L-1/X-287- L-1/X-288- L-1/X-289- L-1/X-290- L-1/X-291-L-1/X-292- L-1/X-293- L-1/X-294- L-1/X-295- L-1/X-296- L-1/X-297-L-1/X-298- L-1/X-299- L-1/X-300- L-1/X-301- L-1/X-302- L-1/X-303-L-1/X-304- L-1/X-305- L-1/X-306- L-1/X-307- L-1/X-308- L-1/X-309-L-1/X-310- L-1/X-311- L-1/X-312- L-1/X-313- L-1/X-314- L-1/X-315-L-1/X-316- L-1/X-317- L-1/X-318- L-1/X-319- L-1/X-320- L-1/X-321-L-1/X-322- L-1/X-323- L-1/X-324- L-1/X-325- L-1/X-326- L-1/X-327-L-1/X-328- L-1/X-329- L-1/X-330- L-1/X-331- L-1/X-332- L-1/X-333-L-1/X-334- L-1/X-335- L-1/X-336- L-1/X-337- L-1/X-338- L-1/X-339-L-1/X-340- L-1/X-341- L-1/X-342- L-1/X-343- L-1/X-344- L-1/X-345-L-1/X-346- L-1/X-347- L-1/X-348- L-1/X-349- L-1/X-350- L-1/X-351-L-1/X-352- L-1/X-353- L-1/X-354- L-1/X-355- L-1/X-356- L-1/X-357-L-1/X-358- L-1/X-359- L-1/X-360- L-1/X-361- L-1/X-362- L-1/X-363-L-1/X-364- L-1/X-365- L-1/X-366- L-1/X-367- L-1/X-368- L-1/X-369-L-1/X-370- L-1/X-371- L-1/X-372- L-1/X-373- L-1/X-374- L-1/X-375-L-1/X-376- L-1/X-377- L-1/X-378- L-1/X-379- L-1/X-380- L-1/X-381-L-1/X-382- L-1/X-383- L-1/X-384- L-1/X-385- L-1/X-386- L-1/X-387-L-1/X-388- L-1/X-389- L-1/X-390- L-1/X-391- L-1/X-392- L-1/X-393-L-1/X-394- L-1/X-395- L-1/X-396- L-1/X-397- L-1/X-398- L-1/X-399-L-1/X-400- L-1/X-401- L-1/X-402- L-1/X-403- L-1/X-404- L-1/X-405-L-1/X-406- L-1/X-407- L-1/X-408- L-1/X-409- L-1/X-410- L-1/X-411-L-1/X-412- L-1/X-413- L-1/X-414- L-1/X-415- L-1/X-416- L-1/X-417-L-1/X-418- L-2/X-1- L-2/X-2- L-2/X-3- L-2/X-4- L-2/X-5- L-2/X-6-L-2/X-7- L-2/X-8- L-2/X-9- L-2/X-10- L-2/X-11- L-2/X-12- L-2/X-13-L-2/X-14- L-2/X-15- L-2/X-16- L-2/X-17- L-2/X-18- L-2/X-19- L-2/X-20-L-2/X-21- L-2/X-22- L-2/X-23- L-2/X-24- L-2/X-25- L-2/X-26- L-2/X-27-L-2/X-28- L-2/X-29- L-2/X-30- L-2/X-31- L-2/X-32- L-2/X-33- L-2/X-34-L-2/X-35- L-2/X-36- L-2/X-37- L-2/X-38- L-2/X-39- L-2/X-40- L-2/X-41-L-2/X-42- L-2/X-43- L-2/X-44- L-2/X-45- L-2/X-46- L-2/X-47- L-2/X-48-L-2/X-49- L-2/X-50- L-2/X-51- L-2/X-52- L-2/X-53- L-2/X-54- L-2/X-55-L-2/X-56- L-2/X-57- L-2/X-58- L-2/X-59- L-2/X-60- L-2/X-61- L-2/X-62-L-2/X-63- L-2/X-64- L-2/X-65- L-2/X-66- L-2/X-67- L-2/X-68- L-2/X-69-L-2/X-70- L-2/X-71- L-2/X-72- L-2/X-73- L-2/X-74- L-2/X-75- L-2/X-76-L-2/X-77- L-2/X-78- L-2/X-79- L-2/X-80- L-2/X-81- L-2/X-82- L-2/X-83-L-2/X-84- L-2/X-85- L-2/X-86- L-2/X-87- L-2/X-88- L-2/X-89- L-2/X-90-L-2/X-91- L-2/X-92- L-2/X-93- L-2/X-94- L-2/X-95- L-2/X-96- L-2/X-97-L-2/X-98- L-2/X-99- L-2/X-100- L-2/X-101- L-2/X-102- L-2/X-103-L-2/X-104- L-2/X-105- L-2/X-106- L-2/X-107- L-2/X-108- L-2/X-109-L-2/X-110- L-2/X-111- L-2/X-112- L-2/X-113- L-2/X-114- L-2/X-115-L-2/X-116- L-2/X-117- L-2/X-118- L-2/X-119- L-2/X-120- L-2/X-121-L-2/X-122- L-2/X-123- L-2/X-124- L-2/X-125- L-2/X-126- L-2/X-127-L-2/X-128- L-2/X-129- L-2/X-130- L-2/X-131- L-2/X-132- L-2/X-133-L-2/X-134- L-2/X-135- L-2/X-136- L-2/X-137- L-2/X-138- L-2/X-139-L-2/X-140- L-2/X-141- L-2/X-142- L-2/X-143- L-2/X-144- L-2/X-145-L-2/X-146- L-2/X-147- L-2/X-148- L-2/X-149- L-2/X-150- L-2/X-151-L-2/X-152- L-2/X-153- L-2/X-154- L-2/X-155- L-2/X-156- L-2/X-157-L-2/X-158- L-2/X-159- L-2/X-160- L-2/X-161- L-2/X-162- L-2/X-163-L-2/X-164- L-2/X-165- L-2/X-166- L-2/X-167- L-2/X-168- L-2/X-169-L-2/X-170- L-2/X-171- L-2/X-172- L-2/X-173- L-2/X-174- L-2/X-175-L-2/X-176- L-2/X-177- L-2/X-178- L-2/X-179- L-2/X-180- L-2/X-181-L-2/X-182- L-2/X-183- L-2/X-184- L-2/X-185- L-2/X-186- L-2/X-187-L-2/X-188- L-2/X-189- L-2/X-190- L-2/X-191- L-2/X-192- L-2/X-193-L-2/X-194- L-2/X-195- L-2/X-196- L-2/X-197- L-2/X-198- L-2/X-199-L-2/X-200- L-2/X-201- L-2/X-202- L-2/X-203- L-2/X-204- L-2/X-205-L-2/X-206- L-2/X-207- L-2/X-208- L-2/X-209- L-2/X-210- L-2/X-211-L-2/X-212- L-2/X-213- L-2/X-214- L-2/X-215- L-2/X-216- L-2/X-217-L-2/X-218- L-2/X-219- L-2/X-220- L-2/X-221- L-2/X-222- L-2/X-223-L-2/X-224- L-2/X-225- L-2/X-226- L-2/X-227- L-2/X-228- L-2/X-229-L-2/X-230- L-2/X-231- L-2/X-232- L-2/X-233- L-2/X-234- L-2/X-235-L-2/X-236- L-2/X-237- L-2/X-238- L-2/X-239- L-2/X-240- L-2/X-241-L-2/X-242- L-2/X-243- L-2/X-244- L-2/X-245- L-2/X-246- L-2/X-247-L-2/X-248- L-2/X-249- L-2/X-250- L-2/X-251- L-2/X-252- L-2/X-253-L-2/X-254- L-2/X-255- L-2/X-256- L-2/X-257- L-2/X-258- L-2/X-259-L-2/X-260- L-2/X-261- L-2/X-262- L-2/X-263- L-2/X-264- L-2/X-265-L-2/X-266- L-2/X-267- L-2/X-268- L-2/X-269- L-2/X-270- L-2/X-271-L-2/X-272- L-2/X-273- L-2/X-274- L-2/X-275- L-2/X-276- L-2/X-277-L-2/X-278- L-2/X-279- L-2/X-280- L-2/X-281- L-2/X-282- L-2/X-283-L-2/X-284- L-2/X-285- L-2/X-286- L-2/X-287- L-2/X-288- L-2/X-289-L-2/X-290- L-2/X-291- L-2/X-292- L-2/X-293- L-2/X-294- L-2/X-295-L-2/X-296- L-2/X-297- L-2/X-298- L-2/X-299- L-2/X-300- L-2/X-301-L-2/X-302- L-2/X-303- L-2/X-304- L-2/X-305- L-2/X-306- L-2/X-307-L-2/X-308- L-2/X-309- L-2/X-310- L-2/X-311- L-2/X-312- L-2/X-313-L-2/X-314- L-2/X-315- L-2/X-316- L-2/X-317- L-2/X-318- L-2/X-319-L-2/X-320- L-2/X-321- L-2/X-322- L-2/X-323- L-2/X-324- L-2/X-325-L-2/X-326- L-2/X-327- L-2/X-328- L-2/X-329- L-2/X-330- L-2/X-331-L-2/X-332- L-2/X-333- L-2/X-334- L-2/X-335- L-2/X-336- L-2/X-337-L-2/X-338- L-2/X-339- L-2/X-340- L-2/X-341- L-2/X-342- L-2/X-343-L-2/X-344- L-2/X-345- L-2/X-346- L-2/X-347- L-2/X-348- L-2/X-349-L-2/X-350- L-2/X-351- L-2/X-352- L-2/X-353- L-2/X-354- L-2/X-355-L-2/X-356- L-2/X-357- L-2/X-358- L-2/X-359- L-2/X-360- L-2/X-361-L-2/X-362- L-2/X-363- L-2/X-364- L-2/X-365- L-2/X-366- L-2/X-367-L-2/X-368- L-2/X-369- L-2/X-370- L-2/X-371- L-2/X-372- L-2/X-373-L-2/X-374- L-2/X-375- L-2/X-376- L-2/X-377- L-2/X-378- L-2/X-379-L-2/X-380- L-2/X-381- L-2/X-382- L-2/X-383- L-2/X-384- L-2/X-385-L-2/X-386- L-2/X-387- L-2/X-388- L-2/X-389- L-2/X-390- L-2/X-391-L-2/X-392- L-2/X-393- L-2/X-394- L-2/X-395- L-2/X-396- L-2/X-397-L-2/X-398- L-2/X-399- L-2/X-400- L-2/X-401- L-2/X-402- L-2/X-403-L-2/X-404- L-2/X-405- L-2/X-406- L-2/X-407- L-2/X-408- L-2/X-409-L-2/X-410- L-2/X-411- L-2/X-412- L-2/X-413- L-2/X-414- L-2/X-415-L-2/X-416- L-2/X-417- L-2/X-418- and so on.and so on.

Utility and Testing

The multibinding compounds of this invention can be used to modulatecalcium channels in various tissues including heart, muscle, andneurons. They will typically be used for the treatment of diseases andconditions in mammals that involve or are mediated by Ca⁺⁺ channels,such as hypertension, cardiac arrythmias, angina pectoris, cerebralischemia, congestive heart failure, migraine, Raynaud's disease, asthmaand bronchospasm, renal impairment and acute renal failure due toprolonged renal ischemia, retinal ischemia, and pain.

The multibinding compounds of this invention are tested in well-knownand reliable assays and their activities are compared with those of thecorresponding unlinked (i.e., monovalent) ligands.

Binding affinity to calcium channels: The binding affinity is determinedby a radioligand competitive displacement assay, essentially asdescribed in Eltze et al, Chirality 2: 233-240 (1990). The ability ofthe present compounds to displace (+)-[³H] isradipine or a similarradioactive ligand from calcium binding sites of guinea pig skeletalmuscle T-tubule membranes is measured in vitro. The binding affinity,calculated from competition curves, is compared with that of themonovalent ligand and/or monovalent linker-ligand conjugate.

Ca⁺⁺ channel activity: The effects of compounds of this invention onchannel activity are determined by measurement of whole-cell Ba⁺⁺currents in voltage-clamped Xenopus oocytes that express various typesof voltage-gated Ca⁺⁺ channels, as described in Bezprozvanny and Tsien,Molec. Pharmacol. 48: 540-549 (1995).

Antivasoconstrictor activity: Antivasoconstrictor activity is determinedas described in Brittain et al, Physiologist 28: 325 (1985) as theconcentration of a compound required to produce 50% vasorelaxation inKCl-contracted rabbit thoracic aorta strips in the presence of calcium.Alternatively, the concentration of a compound required to inhibitcoronary vasoconstriction induced by a thromboxane mimetic (U-46619,i.e., 9,11-methanoepoxy-PGH₂) in guinea pig Langendorff heartpreparation is measured as described in Eltze et al, Chirality 2:233-240 (1990).

Antihypertensive activity: Antihypertensive activity is determined inmale spontaneously hypertensive rats by measurement of mean arterialblood pressure (Rovnyak et al, J. Med. Chem. 35: 3254-3263 (1992)).

Tissue selectivity: Selectivity for vascular smooth muscle as comparedwith cardiac muscle can be assessed by comparing the concentration of amultibinding compound that produces a 50% increase in coronary bloodflow in an isolated guinea-pig heart with that required to inhibitmyocardial contractility. See, e.g., Osterrieder, W and Holck, M., J.Cardiovasc. Pharmacol. 13: 754-9 (1989; and Cremers et al, J.Cardiovasc. Pharmacol. 29: 692-696 (1997).

Pharmaceutical Formulations

When employed as pharmaceuticals, the compounds of Formula I are usuallyadministered in the form of pharmaceutical compositions. This inventiontherefore provides pharmaceutical compositions which contain, as theactive ingredient, one or more of the compounds of Formula I above or apharmaceutically acceptable salt thereof and one or morepharmaceutically acceptable excipients, carriers, diluents, permeationenhancers, solubilizers and adjuvants. The compounds may be administeredalone or in combination with other therapeutic agents (e.g., otherantihypertensive drugs, diuretics and the like). Such compositions areprepared in a manner well known in the pharmaceutical art (see, e.g.,Remington's Pharmaceutical Sciences, Mace Publishing Co., Philadelphia,Pa. 17^(th) Ed. (1985) and “Modem Pharmaceutics”, Marcel Dekker, Inc.3^(rd) Ed. (G. S. Banker & C. T. Rhodes, Eds.).

The compounds of Formula I may be administered by any of the acceptedmodes of administration of agents having similar utilities, for example,by oral, parenteral, rectal, buccal, intranasal or transdermal routes.The most suitable route will depend on the nature and severity of thecondition being treated. Oral administration is a preferred route forthe compounds of this invention. In making the compositions of thisinvention, the active ingredient is usually diluted by an excipient orenclosed within such a carrier which can be in the form of a capsule,sachet, paper or other container. When the excipient serves as adiluent, it can be a solid, semi-solid, or liquid material, which actsas a vehicle, carrier or medium for the active ingredient. Thus, thecompositions can be in the form of tablets, pills, powders, lozenges,sachets, cachets, elixirs, suspensions, emulsions, solutions, syrups,aerosols (as a solid or in a liquid medium), ointments containing, forexample, up to 10% by weight of the active compound, soft and hardgelatin capsules, suppositories, sterile injectable solutions, andsterile packaged powders. Pharmaceutically acceptable salts of theactive agents may be prepared using standard procedures known to thoseskilled in the art of synthetic organic chemistry and described, e.g.,by J. March, Advanced Organic Chemistry: Reactions, Mechanisms andStructure, 4^(th) Ed. (New York: Wiley-Interscience, 1992).

Some examples of suitable excipients include lactose, dextrose, sucrose,sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates,tragacanth, gelatin, calcium silicate, microcrystalline cellulose,polyvinylpyrrolidone, cellulose, sterile water, syrup, and methylcellulose. The formulations can additionally include: lubricating agentssuch as talc, magnesium stearate, and mineral oil; wetting agents;emulsifying and suspending agents; preserving agents such as methyl- andpropylhydroxy-benzoates; sweetening agents; and flavoring agents.

The compositions of the invention can be formulated so as to providequick, sustained or delayed release of the active ingredient afteradministration to the patient by employing procedures known in the art.Controlled release drug delivery systems for oral administration includeosmotic pump systems and dissolutional systems containing polymer-coatedreservoirs or drug-polymer matrix formulations. Examples of controlledrelease systems are given in U.S. Pat. Nos. 3,845,770; 4,326,525;4,902514; and 5,616,345. Another preferred formulation for use in themethods of the present invention employs transdermal delivery devices(“patches”). Such transdermal patches may be used to provide continuousor discontinuous infusion of the compounds of the present invention incontrolled amounts. The construction and use of transdermal patches forthe delivery of pharmaceutical agents is well known in the art. See,e.g., U.S. Pat. Nos. 5,023,252, 4,992,445 and 5,001,139. Such patchesmay be constructed for continuous, pulsatile, or on demand delivery ofpharmaceutical agents.

The compositions are preferably formulated in a unit dosage form. Theterm “unit dosage forms” refers to physically discrete units suitable asunitary dosages for human subjects and other mammals, each unitcontaining a predetermined quantity of active material calculated toproduce the desired therapeutic effect, in association with a suitablepharmaceutical excipient (e.g., a tablet, capsule, ampoule). The activecompound is effective over a wide dosage range and is generallyadministered in a pharmaceutically effective amount. Preferably, fororal administration, each dosage unit contains from 1-250 mg of acompound of Formula I, and for parenteral administration, preferablyfrom 0.1 to 60 mg of a compound of Formula I or a pharmaceuticallyacceptable salt thereof. It will be understood, however, that the amountof the compound actually administered will be determined by a physician,in the light of the relevant circumstances, including the condition tobe treated, the chosen route of administration, the actual compoundadministered and its relative activity, the age, weight, and response ofthe individual patient, the severity of the patient's symptoms, and thelike.

For preparing solid compositions such as tablets, the principal activeingredient is mixed with a pharmaceutical excipient to form a solidpreformulation composition containing a homogeneous mixture of acompound of the present invention. When referring to thesepreformulation compositions as homogeneous, it is meant that the activeingredient is dispersed evenly throughout the composition so that thecomposition may be readily subdivided into equally effective unit dosageforms such as tablets, pills and capsules.

The tablets or pills of the present invention may be coated or otherwisecompounded to provide a dosage form affording the advantage of prolongedaction. For example, the tablet or pill can comprise an inner dosage andan outer dosage component, the latter being in the form of an envelopeover the former. The two components can be separated by an enteric layerwhich serves to resist disintegration in the stomach and permit theinner component to pass intact into the duodenum or to be delayed inrelease. A variety of materials can be used for such enteric layers orcoatings, such materials including a number of polymeric acids andmixtures of polymeric acids with such materials as shellac, cetylalcohol, and cellulose acetate.

The liquid forms in which the novel compositions of the presentinvention may be incorporated for administration orally or by injectioninclude aqueous solutions, suitably flavored syrups, aqueous or oilsuspensions, and flavored emulsions with edible oils such as corn oil,cottonseed oil, sesame oil, coconut oil, or peanut oil, as well aselixirs and similar pharmaceutical vehicles.

Compositions for inhalation or insulation include solutions andsuspensions in pharmaceutically acceptable, aqueous or organic solvents,or mixtures thereof, and powders. The liquid or solid compositions maycontain suitable pharmaceutically acceptable excipients as describedsupra. Preferably the compositions are administered by the oral or nasalrespiratory route for local or systemic effect. Compositions inpreferably pharmaceutically acceptable solvents may be nebulized by useof inert gases. Nebulized solutions may be inhaled directly from thenebulizing device or the nebulizing device may be attached to a facemask tent, or intermittent positive pressure breathing machine.Solution, suspension, or powder compositions may be administered,preferably orally or nasally, from devices which deliver the formulationin an appropriate manner.

The following formulation examples illustrate representativepharmaceutical compositions of the present invention.

FORMULATION EXAMPLE 1

Hard gelatin capsules containing the following ingredients are prepared:

Quantity Ingredient (mg/capsule) Active Ingredient 30.0 Starch 305.0Magnesium stearate 5.0

The above ingredients are mixed and filled into hard gelatin capsules in340 mg quantities.

FORMULATION EXAMPLE 2

A tablet formula is prepared using the ingredients below:

Quantity Ingredient (mg/tablet) Active Ingredient 25.0 Cellulose,microcrystalline 200.0 Colloidal silicon dioxide 10.0 Stearic acid 5.0

The components are blended and compressed to form tablets, each weighing240 mg.

FORMULATION EXAMPLE 3

A dry powder inhaler formulation is prepared containing the followingcomponents:

Ingredient Weight % Active Ingredient 5 Lactose 95

The active ingredient is mixed with the lactose and the mixture is addedto a dry powder inhaling appliance.

FORMULATION EXAMPLE 4

Tablets, each containing 30 mg of active ingredient, are prepared asfollows:

Quantity Ingredient (mg/tablet) Active Ingredient 30.0 mg Starch 45.0 mgMicrocrystalline cellulose 35.0 mg Polyvinylpyrrolidone 4.0 mg (as 10%solution in sterile water) Sodium carboxymethyl starch 4.5 mg Magnesiumstearate 0.5 mg Talc 1.0 mg Total 120 mg

The active ingredient, starch and cellulose are passed through a No. 20mesh U.S. sieve and mixed thoroughly. The solution ofpolyvinylpyrrolidone is mixed with the resultant powders, which are thenpassed through a 16 mesh U.S. sieve. The granules so produced are driedat 50° C. to 60° C. and passed through a 16 mesh U.S. sieve. The sodiumcarboxymethyl starch, magnesium stearate, and talc, previously passedthrough a No. 30 mesh U.S. sieve, are then added to the granules which,after mixing, are compressed on a tablet machine to yield tablets eachweighing 120 mg.

FORMULATION EXAMPLE 5

Capsules, each containing 40 mg of medicament are made as follows:

Quantity Ingredient (mg/capsule) Active Ingredient 40.0 mg Starch 109.0mg Magnesium stearate 1.0 mg Total 150.0 mg

The active ingredient, starch, and magnesium stearate are blended,passed through a No. 20 mesh U.S. sieve, and filled into hard gelatincapsules in 150 mg quantities.

FORMULATION EXAMPLE 6

Suppositories, each containing 25 mg of active ingredient are made asfollows:

Ingredient Amount Active Ingredient 25 mg Saturated fatty acidglycerides to 2,000 mg

The active ingredient is passed through a No. 60 mesh U.S. sieve andsuspended in the saturated fatty acid glycerides previously melted usingthe minimum heat necessary. The mixture is then poured into asuppository mold of nominal 2.0 g capacity and allowed to cool.

FORMULATION EXAMPLE 7

Suspensions, each containing 50 mg of medicament per 5.0 mL dose aremade as follows:

Ingredient Amount Active Ingredient 50.0 mg Xanthan gum 4.0 mg Sodiumcarboxymethyl cellulose (11%) Microcrystalline cellulose (89%) 50.0 mgSucrose 1.75 g Sodium benzoate 10.0 mg Flavor and Color q.v. Purifiedwater to 5.0 mL

The active ingredient, sucrose and xanthan gum are blended, passedthrough a No. 10 mesh U.S. sieve, and then mixed with a previously madesolution of the microcrystalline cellulose and sodium carboxymethylcellulose in water. The sodium benzoate, flavor, and color are dilutedwith some of the water and added with stirring. Sufficient water is thenadded to produce the required volume.

FORMULATION EXAMPLE 8

Quantity Ingredient (mg/capsule) Active Ingredient 15.0 mg Starch 407.0mg Magnesium stearate 3.0 mg Total 425.0 mg

The active ingredient, starch, and magnesium stearate are blended,passed through a No. 20 mesh U.S. sieve, and filled into hard gelatincapsules in 425.0 mg quantities.

FORMULATION EXAMPLE 9

A subcutaneous formulation may be prepared as follows:

Ingredient Quantity Active Ingredient 5.0 mg Corn Oil 1.0 mL

Frequently, it will be desirable or necessary to introduce thepharmaceutical composition to the brain, either directly or indirectly.Direct techniques usually involve placement of a drug delivery catheterinto the host's ventricular system to bypass the blood-brain barrier.One such implantable delivery system used for the transport ofbiological factors to specific anatomical regions of the body isdescribed in U.S. Pat. No. 5,011,472 which is herein incorporated byreference.

Indirect techniques, which are generally preferred, usually involveformulating the compositions to provide for drug latentiation by theconversion of hydrophilic drugs into lipid-soluble drugs. Latentiationis generally achieved through blocking of the hydroxy, carbonyl,sulfate, and primary amine groups present on the drug to render the drugmore lipid soluble and amenable to transportation across the blood-brainbarrier. Alternatively, the delivery of hydrophilic drugs may beenhanced by intra-arterial infusion of hypertonic solutions which cantransiently open the blood-brain barrier.

SYNTHETIC EXAMPLES Example 1 (See FIG. 4)

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the amlodipine moiety linked via the side chain amine tothe linker, X.

Method A

Step 1

A solution of N-BOC-amlodipine [structure 9, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol), the linker molecule 1,6-dibromohexane (1 mmol),and diisopropylethylamine (0.2 mL) in DMF (3 mL) is stirred and warmedunder an inert atmosphere. The progress of the reaction is followed bytlc and when reaction is complete, the solution is poured into aqueous5% NaHCO₃ and the aqueous mixture is extracted with methylene chloride.The organic extract solution is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product by useof HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 11, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-amlodipine in the above examplewith other ligands of structure 9 and/or by replacing 1,6-dibromohexanewith other linker molecules, other compounds of Formula I are prepared.

Method B

A solution of amlodipine [structure 9, where PG is H; R⁶ and R⁸ aremethyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol), the linkermolecule 1,6-dibromohexane (1 mmol), and diisopropylethylamine (0.2 mL)in DMF (3 mL) is stirred and warmed under an inert atmosphere. Theprogress of the reaction is followed by tlc and when reaction iscomplete, the solution is poured into aqueous 5% NaHCO₃ and the aqueousmixture is extracted with methylene chloride. The organic extractsolution is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound of Formula I(structure 11, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; andR¹⁰ is H) is obtained by purification of the crude product by use ofHPLC.

In similar manner, by replacing amlodipine in the above example withother ligands of structure 9 and/or by replacing 1,6-dibromohexane withother linker molecules, other compounds of Formula I are prepared.

Example 2 See FIG. 4

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the amlodipine moiety linked via the side chain amine tothe linker, X, through an amide bond

Method A

Step 1

A solution of N-BOC-amlodipine [structure 9, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol), the linker molecule 3,6-dioxaoctanedioic acid (1mmol) in CH₂Cl₂ (5 mL) is prepared under argon in a flask equipped withmagnetic stirrer and a drying tube. To this solution is addeddicyclohexylcarbodiimide (solid, 2.2 mmol). The progress of the reactionis followed by tlc and after reaction occurs, the reaction solution isquenched in water, aqueous sodium bicarbonate is added and the aqueousmixture is extracted with methylene chloride. The organic layer iswashed with aqueous Na₂CO₃ and with H₂O, dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product withthe use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 14, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-amlodipine in the above examplewith other ligands of structure 9 and/or by replacing3,6-dioxaoctanedioic acid with other linker molecules, other compoundsof Formula I are prepared.

Method B

A solution of amlodipine [structure 9, where PG is H; R⁶ and R⁸ aremethyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol), the linkermolecule 3,6-dioxaoctanedioic acid (1 mmol) in CH₂Cl₂ (5 mL) is preparedunder argon in a flask equipped with magnetic stirrer and a drying tube.To this solution is added dicyclohexylcarbodiimide (solid, 2.2 mmol).The progress of the reaction is followed by tlc and after reactionoccurs, the reaction solution is quenched in water, aqueous sodiumbicarbonate is added and the aqueous mixture is extracted with methylenechloride. The organic layer is washed with aqueous Na₂CO₃ and with H₂O,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired Formula I compound (structure 14, whereR⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtainedby purification of the crude product with the use of HPLC.

In similar manner, by replacing amlodipine in the above example withother ligands of structure 9 and/or by replacing 3,6-dioxaoctanedioicacid with other linker molecules, other compounds of Formula I areprepared.

Example 3 See FIG. 4

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the amlodipine moiety linked via the side chain amine tothe linker, X, through a urea bond

Method A

Step 1

A solution of the linker molecule 1,4-diisocyanatobutane (1 mmol) inCH₂Cl₂ (5 mL) containing Et₃N (0.2 mL) is stirred and cooled in anice-water bath under an inert atmosphere. To this is added dropwise asolution of N-BOC-amlodipine [structure 9, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5 mL). After addition is complete, thecooling bath is removed and the reaction solution is allowed to warm toroom temperature. The progress of the reaction is followed by tlc andwhen reaction has occurred, the reaction solution is quenched in cold 5%aqueous Na₂CO₃. The layers are separated and the organic layer is washedwith aqueous Na₂CO₃, with water and is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product withthe use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 16, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-amlodipine in the above examplewith other ligands of structure 9 and/or by replacing1,4-diisocyanatobutane with other linker molecules, other compounds ofFormula I are prepared.

Method B

A solution of the linker molecule 1,4-diisocyanatobutane (1 mmol) inCH₂Cl₂ (5 mL) containing Et₃N (0.2 mL) is stirred and cooled in anice-water bath under an inert atmosphere. To this is added dropwise asolution of amlodipine [structure 9, where PG is H; R⁶ and R⁸ aremethyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5mL). After addition is complete, the cooling bath is removed and thereaction solution is allowed to warm to room temperature. The progressof the reaction is followed by tlc and when reaction has occurred, thereaction solution is quenched in cold 5% aqueous Na₂CO₃. The layers areseparated and the organic layer is washed with aqueous Na₂CO₃, withwater and is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired Formula I compound(structure 16, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; andR¹⁰ is H) is obtained by purification of the crude product with the useof HPLC.

In similar manner, by replacing amlodipine in the above example withother ligands of structure 9 and/or by replacing 1,4-diisocyanatobutanewith other linker molecules, other compounds of Formula I are prepared.

Example 4 See FIG. 4

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the amlodipine moiety linked via the side chain amine tothe linker, X, through a sulfonamide bond.

Method A

Step 1

A solution of the linker molecule benzene-1,4-bis-sulfonyl chloride (1mmol) in CH₂Cl₂(5 mL) containing Et₃N (0.2 mL) is stirred and cooled inan ice-water bath under an inert atmosphere. To this is added dropwise asolution of N-BOC-amlodipine [structure 9, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5 mL). After addition is complete, thecooling bath is removed and the reaction solution is allowed to warm toroom temperature. The progress of the reaction is followed by tlc andwhen reaction has occurred, the reaction solution is quenched in cold 5%aqueous Na₂CO₃. The layers are separated and the organic layer is washedwith aqueous Na₂CO₃, with water and is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product withthe use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 18, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-amlodipine in the above examplewith other ligands of structure 9 and/or by replacingbenzene-1,4-bissulfonyl chloride with other linker molecules, othercompounds of Formula I are prepared.

Method B

A solution of the linker molecule benzene-1,4-bissulfonyl chloride (1mmol) in CH₂Cl₂ (5 mL) containing Et₃N (0.2 mL) is stirred and cooled inan ice-water bath under an inert atmosphere. To this is added dropwise asolution of amlodipine [structure 9, where PG is H; R⁶ and R⁸ aremethyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5mL). After addition is complete, the cooling bath is removed and thereaction solution is allowed to warm to room temperature. The progressof the reaction is followed by tlc and when reaction has occurred, thereaction solution is quenched in cold 5% aqueous Na₂CO₃. The layers areseparated and the organic layer is washed with aqueous Na₂CO₃, withwater and is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired Formula I compound(structure 18, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; andR¹⁰ is H) is obtained by purification of the crude product with the useof HPLC.

In similar manner, by replacing amlodipine in the above example withother ligands of structure 9 and/or by replacing benzene-1,4-bissulfonylchloride with other linker molecules, other compounds of Formula I areprepared.

Example 5 See FIG. 5

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the 1,4-dihydropyridine moiety linked via the2-hydroxymethyl group to the linker, X, through an ether bond.

Method A

Step 1

To a mixture of sodium hydride (3.1 mmol) and dry THF (2 mL) stirredunder an inert atmosphere and protected from the atmosphere with adrying tube is added a solution of the linker molecule1,4-dihydroxymethylbenzene (1 mmol) in dry THF (2 mL). To this is addeda solution of N-BOC-1,4-dihydropyridine [structure 25, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol) in dry THF (2 mL). A solution of is added and theresulting mixture is stirred at RT. The progress of the reaction isfollowed by Uc and when reaction is complete, the solution is pouredinto aqueous 5% NaHCO₃ and the aqueous mixture is extracted withmethylene chloride. The organic extract solution is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound is obtained by purification of the crudeproduct by use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 22, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-1,4-dihydropyridine in the aboveexample with other ligands of structure 21 and/or by replacing1,4-dihydroxymethylbenzene-with other linker molecules, other compoundsof Formula I are prepared.

Method B

Step 1

To a mixture of sodium hydride (3.1 mmol) and dry THF (2 mL) stirredunder an inert atmosphere and protected from the atmosphere with adrying tube is added a solution of the linker molecule1,4-dihydroxymethylbenzene (1 mmol) in dry THF (2 mL). To this is addeda solution of 1,4-dihydropyridine [structure 25, where PG is H; R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol) in dryTHF (2 mL). A solution of is added and the resulting mixture is stirredat RT. The progress of the reaction is followed by tlc and when reactionis complete, the solution is poured into aqueous 5% NaHCO₃ and theaqueous mixture is extracted with methylene chloride. The organicextract solution is dried (Na₂SO₄), filtered and concentrated underreduced pressure to give the crude product. The desired compound ofFormula I (structure 22, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is2-Cl; and R¹⁰ is H) is obtained by purification of the crude product byuse of HPLC.

In similar manner, by replacing 1,4-dihydropyridine in the above examplewith other ligands of structure 25 and/or by replacing1,4-dihydroxymethylbenzene with other linker molecules, other compoundsof Formula I are prepared.

Example 6 See FIG. 5

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the 1,4-dihydropyridine moiety linked via the2-hydroxymethyl group to the linker, X, through an ester bond

Method A

Step 1

A solution of N-BOC-1,4dihydropyridine [structure 21, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R is H] (2 mmol), the linker molecule benzene-1,4-bisacetic acid (1mmol), and 4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is preparedunder argon in a flask equipped with magnetic stirrer and a drying tube.To this solution is added dicyclohexylcarbodiimide (solid, 2.2 mmol).The progress of the reaction is followed by tlc and after reactionoccurs, the reaction solution is quenched in water, aqueous sodiumbicarbonate is added and the aqueous mixture is extracted with methylenechloride. The organic layer is washed with aqueous Na₂CO₃ and with H₂O,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound is obtained by purification ofthe crude product with the use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 23, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-1,4-dihydropyridine in the aboveexample with other ligands of structure 21 and/or by replacingbenzene-1,4-bisacetic acid with other linker molecules, other compoundsof Formula I are prepared.

Method B

A solution of 1,4-dihydropyridine [structure 21, where PG is H; R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol), thelinker molecule benzene-1,4-bisacetic acid (1 mmol), and4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is prepared under argonin a flask equipped with magnetic stirrer and a drying tube. To thissolution is added dicyclohexylcarbodiimide (solid, 2.2 mmol). Theprogress of the reaction is followed by tlc and after reaction occurs,the reaction solution is quenched in water, aqueous sodium bicarbonateis added and the aqueous mixture is extracted with methylene chloride.The organic layer is washed with aqueous Na₂CO₃ and with H₂O, dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound of Formula I (structure 23, where R⁶and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing 1,4-dihydropyridine in the above examplewith other ligands of structure 21 and/or by replacingbenzene-1,4-bisacetic acid with other linker molecules, other compoundsof Formula I are prepared.

Example 7 See FIG. 5

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the 1,4-dihydropyridine moiety linked via the2-hydroxymethyl group to the linker, X, through a carbamate bond

Method A

Step 1

A solution of the linker molecule trans-1,4-cyclohexylisocyanate (1mmol) in CH₂Cl₂ (5 mL) containing Et₃N (0.2 mL) is stirred and cooled inan ice-water bath under an inert atmosphere. To this is added dropwise asolution of N-BOC-1,4dihydropyridine [structure 21, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5 mL). After addition is complete, thecooling bath is removed and the reaction solution is allowed to warm toroom temperature. The progress of the reaction is followed by tlc andwhen reaction has occurred, the reaction solution is quenched in cold 5%aqueous Na₂CO₃. The layers are separated and the organic layer is washedwith aqueous Na₂CO₃, with water and is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product withthe use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 24, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-1,4-dihydropyridine in the aboveexample with other ligands of structure 21 and/or by replacingtrans-1,4-cyclohexylisocyanate with other linker molecules, othercompounds of Formula I are prepared.

Method B

A solution of the linker molecule trans-1,4cyclohexylisocyanate (1 mmol)in CH₂Cl₂(5 mL) containing Et₃N (0.2 mL) is stirred and cooled in anice-water bath under an inert atmosphere. To this is added dropwise asolution of1,4-dihydropyridine [structure 21, where PG is H; R⁶ and R⁸are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol) in CH₂Cl₂ (5mL). After addition is complete, the cooling bath is removed and thereaction solution is allowed to warm to room temperature. The progressof the reaction is followed by tlc and when reaction has occurred, thereaction solution is quenched in cold 5% aqueous Na₂CO₃. The layers areseparated and the organic layer is washed with aqueous Na₂CO₃, withwater and is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound of Formula I(structure 24, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; andR¹⁰ is H) is obtained by purification of the crude product with the useof HPLC.

In similar manner, by replacing 1,4-dihydropyridine in the above examplewith other ligands of structure 21 and/or by replacingtrans-1,4-cyclohexylisocyanate with other linker molecules, othercompounds of Formula I are prepared.

Example 8 See FIG. 8

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the 1,4-dihydropyridine moiety linked via the 3-carboxylgroup to the linker, X, through an ester bond

Method A

Step 1

A solution of N-BOC-1,4-dihydropyridine (structure 36, where PG ist-butyloxycarbonyl (BOC); R², R⁶ and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰is H] (2 mmol), the linker molecule 1,4-bis(hydroxymethyl)benzene (1mmol), and 4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is preparedunder argon in a flask equipped with magnetic stirrer and a drying tube.To this solution is added dicyclohexylcarbodiimide (solid, 2.2 mmol).The progress of the ms reaction is followed by tlc and after reactionoccurs, the reaction solution is quenched in water, aqueous sodiumbicarbonate is added and the aqueous mixture is extracted with methylenechloride. The organic layer is washed with aqueous Na₂CO₃ and with H₂O,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound is obtained by purification ofthe crude product with the use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 37, where R², R⁶and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained by purificationof the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-1,4-dihydropyridine in the aboveexample with other ligands of structure 36 and/or by replacing1,4-bis(hydroxymethyl)benzene with other linker molecules, othercompounds of Formula I are prepared.

Method B

A solution of dihydropyridine (structure 36, where PG is H; R², R⁶ andR⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol), the linker molecule1,4-bis(hydroxymethyl)benzene (1 mmol), and 4-dimethylaminopyridine (10mg) in CH₂Cl₂ (5 mL) is prepared under argon in a flask equipped withmagnetic stirrer and a drying tube. To this solution is addeddicyclohexylcarbodiimide (solid, 2.2 mmol). The progress of the reactionis followed by tlc and after reaction occurs, the reaction solution isquenched in water, aqueous sodium bicarbonate is added and the aqueousmixture is extracted with methylene chloride. The organic layer iswashed with aqueous Na₂CO₃ and with H₂O, dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound of Formula I (structure 37, where R², R⁶ and R⁸ aremethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained by purification of thecrude product with the use of HPLC.

In similar manner, by replacing 1,4-dihydropyridine in the above examplewith other ligands of structure 36 and/or by replacing1,4-bis(hydroxymethyl)benzene with other linker molecules, othercompounds of Formula I are prepared.

Example 9 See FIG. 8

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the 1,4-dihydropyridine moiety linked via the 3-carboxylgroup to the linker, X, through an amide bond.

Method A

Step 1

A solution of N-BOC-1,4-dihydropyridine (structure 36, where PG ist-butyloxycarbonyl (BOC); R², R⁶ and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰is H] (2 mmol), the linker molecule 1,5diaminopentane (1 mmol), and4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is prepared under argonin a flask equipped with magnetic stirrer and a drying tube. To thissolution is added dicyclohexylcarbodiimide (solid, 2.2 mmol). Theprogress of the reaction is followed by Uc and after reaction occurs,the reaction solution is quenched in water, aqueous sodium bicarbonateis added and the aqueous mixture is extracted with methylene chloride.The organic layer is washed with aqueous Na₂CO₃ and with H₂O, dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound is obtained by purification of thecrude product with the use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by Uc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I (structure 37a, where R², R⁶and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained by purificationof the crude product with the use of HPLC.

In similar manner, by replacing N-BOC-1,4-dihydropyridine in the aboveexample with other ligands of structure 36 and/or by replacing1,5-diaminopentane with other linker molecules, other compounds ofFormula I are prepared.

Method B

A solution of dihydropyridine (structure 36, where PG is H; R², R⁶ andR⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H] (2 mmol), the linker molecule1,5-diaminopentane (1 mmol), and 4-dimethylaminopyridine (10 mg) inCH₂Cl₂ (5 mL) is prepared under argon in a flask equipped with magneticstirrer and a drying tube. To this solution is addeddicyclohexylcarbodiimide (solid, 2.2 mmol). The progress of the reactionis followed by tlc and after reaction occurs, the reaction solution isquenched in water, aqueous sodium bicarbonate is added and the aqueousmixture is extracted with methylene chloride. The organic layer iswashed with aqueous Na₂CO₃ and with H₂O, dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired Formula I compound (structure 37a, where R², R⁶ and R⁸ aremethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained by purification of thecrude product with the use of HPLC.

In similar manner, by replacing dihydropyridine in the above examplewith other ligands of structure 36 and/or by replacing1,5-diaminopentane with other linker molecules, other compounds ofFormula I are prepared.

Example 10 See FIG. 10

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the benzothiazepine moiety linked via the nitrogen of theamide group to the linker, X

A mixture of NaH (2.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added first a solution of benzothiazepine (structure 41, whereR¹² is OAc; R¹³ is Me; and R¹⁴ is H) (2 mmol) in DMF (5 mL) and then thelinker molecule 1,8-dibromooctane (1 mmol). The resulting mixture isstirred and the course of the reaction is followed by thin layerchromatography. After reaction occurs, the reaction is quenched withcold dilute aq. Na₂CO₃ and extracted with methylene chloride. Theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound of Formula I(structure 42, where R¹² is OAc; R¹³ is Me; and R¹⁴ is H) is obtained bypurification of the crude product by use of HPLC.

In similar manner, by replacing the benzothiazepine in the above examplewith other ligands of structure 41 and/or by replacing 1,8-dibromooctanewith other linker molecules, other compounds of Formula I are prepared.

Example 11 See FIG. 10

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the benzothiazepine moiety linked via the 3-hydroxyl groupto the linker, X.

Step 1

A solution of the benzothiazepine (structure 43, where PG=Ac; R¹¹ is2-(N,N-dimethylamino)ethyl; R¹³ is methyl; and R¹⁴ is H) (1 mmol) inmethanol (5 mL) is stirred with potassium carbonate. The progress of thereaction is followed by tlc. After reaction occurs, the mixture isfiltered to remove solids and the filtrate is concentrated to give thecrude product. The desired compound (structure 44) is obtained bypurification of the crude product with the use of HPLC.

Step 2

A solution of the linker molecule benzene-1,4-bisacetyl chloride (2mmol) in methylene chloride is added slowly to a solution of thebenzothiazepine [structure 44, where R¹¹ is 2-(N,N-dimethylamino)ethyl;R¹³ is methyl; and R¹⁴ is H] (2 mmols) in methylene chloride (5 mL) andpyridine (0.5 mL) in a flask equipped with a magnetic stirrer and adrying tube and which is cooled in an ice-water bath. The course of thereaction is followed by thin layer chromatography. After reactionoccurs, the reaction solution is quenched in water and the aqueousmixture is extracted with ethyl acetate. The organic layer is washedwith aqueous Na₂CO₃ with water, and is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound of Formula I [structure 45, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹³ is methyl; and R¹⁴ is H] is obtained bypurification of the crude product by use of HPLC.

In similar manner, by replacing the benzothiazepine in the above examplewith other ligands of structure 44 and/or by replacingbenzene-1,4-bisacetyl chloride with other linker molecules, othercompounds of Formula I are prepared.

Example 12 See FIG. 10

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the benzothiazepine moiety linked via the oxygen of thephenolic group to the linker, X

Step 1

A solution of the benzothiazepine [structure 77, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] (2 mmols) inmethylene chloride (5 mL) is stirred and cooled to −78° C. under andinert atmosphere. BBr₃ (5 mmol) is added and stirring is continued asthe cooling bath is removed and the temperature of the reaction solutionis allowed to rise to room temperature. The course of the reaction isfollowed by thin layer chromatography. After reaction occurs, thereaction solution is diluted with methylene chloride and washed withcold aqueous Na₂CO₃ and with half-saturated brine. The organic layer isdried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound [structure 78, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] is obtained bypurification of the crude product by use of HPLC.

Step 2

A solution of the benzothiazepine [structure 78, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] (2 mmols) andlinker molecule 1,4bisiodomethylbenzene (1 mmol) in acetone (5 mL)containing K₂CO₃ is stirred and heated at reflux temperature under aninert atmosphere. The course of the reaction is followed by thin layerchromatography. After reaction occurs, the reaction solution is dilutedwith ethyl acetate and washed with water and with aqueous Na₂CO₃. Theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound of Formula I[structure 79, where R¹¹ is 2-(N,N-dimethylamino)ethyl; R¹² is OAc; andR¹⁴ is H] is obtained by purification of the crude product by use ofHPLC.

In similar manner, by replacing the benzothiazepine in the above examplewith other ligands of structure 78 and/or by replacing1,4-bisiodomethylbenzene with other linker molecules, other compounds ofFormula I are prepared.

Example 13 See FIG. 11

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the verapamil moiety linked via the nitrogen of the aminegroup to the linker, X.

A solution of N-desmethyl-verapamil (structure 82) (2 mmol), the linkermolecule 1,4-diiodobutane (1 mmol), and diisopropylethylamine (0.2 mL)in DMF (3 mL) is stirred and warmed under an inert atmosphere. Theprogress of the reaction is followed by tlc and when reaction iscomplete, the solution is poured into aqueous 5% NaHCO₃ and the aqueousmixture is extracted with methylene chloride. The organic extractsolution is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound of Formula I(structure 83) is obtained by purification of the crude product by useof HPLC.

Example 14 See FIG. 11

Preparation of a Formula I compound wherein p is 2, q is 1, and theligand, L, is the verapamil moiety linked via an oxygen atom to thelinker, X.

A solution of the O-desmethyl-verapamil (structure 85) (2 mmols) andlinker molecule 1,2-bis-(2-iodoethoxy)ethane (1 mmol) in acetone (5 mL)containing K₂CO₃ is stirred and heated at reflux temperature under aninert atmosphere. The course of the reaction is followed by thin layerchromatography. After reaction occurs, the reaction solution is dilutedwith ethyl acetate and washed with water and with aqueous Na₂CO₃. Theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired Formula I compound(structure 86) is obtained by purification of the crude product by useof HPLC.

Example 15 See FIG. 16

Preparation of a Formula I compound wherein p is 2, q is 1, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via the 3-carboxylgroup to the linker, X, and the second ligand, L₂, is thebenzothioazepine moiety linked to linker, X, via the hydroxyl functionof the phenolic ring.

Method A

Step 1

A solution of the benzothiazepine [structure 78, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H (see FIG. 13)] (1mmols) and linker molecule 1-iodomethyl-4-benzyloxybenzene (1 mmol) inacetone (5 mL) containing K₂CO₃ is stirred and heated at refluxtemperature under an inert atmosphere. The course of the reaction isfollowed by thin layer chromatography. After reaction occurs, thereaction solution is diluted with ethyl acetate and washed with waterand with aqueous Na₂CO₃. The organic layer is dried (Na₂SO₄), filteredand concentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product by useof HPLC.

Step 2

Ammonium formate (96 mg, 1.5 mmol) and 10% Pd—C (50 mg) are added to asolution of the compound obtained in the preceding reaction in methanol(3 mL) and THF (2 mL). The mixture is stirred at room temperature andthe progress of the reaction is monitored by tlc. After reaction iscomplete, the mixture is filtered through Celite, the filter pad isrinsed with EtOAc, the combined organic layers are washed successivelywith aq. NaHCO₃ and with half-saturated brine, then filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound [(see FIG. 13 and 16) structure 52, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] is obtained bypurification of the crude product with the use of HPLC.

Step 3

A solution of N-BOC-1,4-dihydropyridine (structure 36, where PG ist-butyloxycarbonyl (BOC); R² is 2-(N-BOC-amino)ethoxymethyl; R⁶ and R⁸are methyl; R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol), the compound [structure52, where R¹¹ is 2-(N,N-dimethylamino)ethyl; R¹² is Oac; and R¹⁴ is H](1 mmol) obtained in the preceding reaction (1 mmol), and4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is prepared under argonin a flask equipped with magnetic stirrer and a drying tube. To thissolution is added dicyclohexylcarbodiimide (solid, 2.2 mmol). Theprogress of the reaction is followed by tlc and after reaction occurs,the reaction solution is quenched in water, aqueous sodium bicarbonateis added and the aqueous mixture is extracted with methylene chloride.The organic layer is washed with aqueous Na₂CO₃ and with H₂O, dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound is obtained by purification of thecrude product with the use of HPLC.

Step 4

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 68, where R² is(2-amino)ethoxymethyl; R⁶ and R⁸ are methyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ areH; R¹¹ is 2-(N,N-dimethylamino)ethyl; and R¹² is OAc] is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing the 1,4dihydropyridine of the aboveexample with other ligands of structure 36 and/or the benzothiazepine inthe above example with other ligands of structure 52 and/or by replacing1-iodomethyl-4-benzyloxybenzene with other linker molecules, othercompounds of Formula I are prepared.

Method B

Step 1

A solution of dihydropyridine (structure 36, where PG is H; R² is2-(N-BOC-amino)ethoxymethyl; R⁶ and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰is H] (1 mmol), the compound [structure 52, where R¹¹ is2-(N,N-dimethylamino) ethyl; R¹² is OAc; and R¹⁴ is H] (1 mmol) obtainedin the preceding reaction (1 mmol), and 4-dimethylaminopyridine (10 mg)in CH₂Cl₂ (5 mL) is prepared under argon in a flask equipped withmagnetic stirrer and a drying tube. To this solution is addeddicyclohexylcarbodiimide (solid, 2.2 mmol). The progress of the reactionis followed by tlc and after reaction occurs, the reaction solution isquenched in water, aqueous sodium bicarbonate is added and the aqueousmixture is extracted with methylene chloride. The organic layer iswashed with aqueous Na₂CO₃ and with H₂O, dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product withthe use of HPLC.

Step 2

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired Formula I compound [structure 68, where R² is(2-amino)ethoxymethyl; R⁶ and R⁸ are methyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ areH; R¹¹ is 2-(N,N-dimethylamino)ethyl; and R¹² is OAc] is obtained bypurification of the crude product with the use of HPLC.

Example 16 See FIG. 16

Preparation of a Formula I compound wherein p is 2, q is 1, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via the2-hydroxymethyl group to the linker, X, and the second ligand, L₂, isthe benzothioazepine moiety linked to X via the hydroxyl function of thephenolic ring

Method A

Step 1

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the linker molecule1-hydroxymethyl-4-benzyloxybenzene (1 mmol) in dry DMF (5 mL) and theresulting mixture is stirred for 1 hour. Then a solution of theN-BOC-1,4-dihydropyridine [(see FIG. 12) structure 21, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] (1 mmol) in dry DMF (2 mL) is added. The resulting mixtureis stirred and the course of the reaction is followed by thin layerchromatography. After reaction occurs, the reaction is quenched withcold dilute aq. Na₂CO₃ and extracted with methylene chloride. Theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound is obtained bypurification of the crude product by use of HPLC.

Step 2

Ammonium formate (96 mg, 1.5 mmol) and 10% Pd—C (50 mg) are added to asolution of the compound obtained in the preceding reaction in methanol(3 mL) and THF (2 mL). The mixture is stirred at room temperature andthe progress of the reaction is monitored by tlc. After reaction iscomplete, the mixture is filtered through Celite, the filter pad isrinsed with EtOAc, the combined organic layers are washed successivelywith aq. NaHCO₃ and with half-saturated brine, then filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound [(see FIG. 12 and 16) structure 47, where PG ist-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H] is obtained by purification of the crude product with theuse of HPLC.

Step 3

Diethyl azodicarboxylate (1 mmol) is added dropwise via a syringe to astirred solution of triphenylphosphine (1 mmol) in THF (5 mL) at roomtemperature. To this is added a solution of the compound obtained in thepreceding reaction (structure 47, where PG is t-butyloxycarbonyl (BOC);R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) (1 mmol)and the benzothiazepine [structure 78, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] (1 mmol) in THF (3mL). The resulting solution is stirred at RT and the progress of thereaction is followed by tlc. After reaction occurs, solvent is removedby evaporation under reduced pressure and the residue is purified byHPLC, giving the desired compound.

Step 4

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 69, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H: R¹¹ is2-(N,N-dimethylamino)ethyl; and R¹² is OAc] is obtained by purificationof the crude product with the use of HPLC.

In similar manner, by replacing the 1,4dihydropyridine of the aboveexample with other ligands of structure 47 and/or the benzothiazepine inthe above example with other ligands of structure 78 and/or by replacing1-bromomethyl-4-benzyloxybenzene with other linker molecules, othercompounds of Formula I are prepared.

Method B

Step 1

A mixture of NaH (2.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the linker molecule1-hydroxymethyl-4-benzyloxybenzene (1 mmol) in dry DMF (5 mL) and theresulting mixture is stirred for 1 hour. Then a solution of the1,4-dihydropyridine [(see FIG. 12) structure 21, where PG is H; R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol) in dryDMF (2 mL) is added. The resulting mixture is stirred and the course ofthe reaction is followed by thin layer chromatography. After reactionoccurs, the reaction is quenched with cold dilute aq. Na₂CO₃ andextracted with methylene chloride. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound is obtained by purification of the crudeproduct by use of HPLC.

Step 2

Ammonium formate (96 mg, 1.5 mmol) and 10% Pd—C (50 mg) are added to asolution of the compound obtained in the preceding reaction in methanol(3 mL) and THF (2 mL). The mixture is stirred at room temperature andthe progress of the reaction is monitored by tlc. After reaction iscomplete, the mixture is filtered through Celite, the filter pad isrinsed with EtOAc, the combined organic layers are washed successivelywith aq. NaHCO₃ and with half-saturated brine, then filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound [(see FIGS. 12 and 16) structure 47, where PG is H; R⁶and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] is obtained bypurification of the crude product with the use of HPLC.

Step 3

Diethyl azodicarboxylate (1 mmol) is added dropwise via a syringe to astirred solution of triphenylphosphine (1 mmol) in THF (5 mL) at roomtemperature. To this is added a solution of the compound obtained in thepreceding reaction (structure 47, where PG is H; R6 and R⁸ are methyl;R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) (1 mmol) and the benzothiazepine[structure 78, where R¹¹ is 2-(N,N-dimethylamino)ethyl; R¹² is OAc; andR¹⁴ is H] (1 mmol) in THF (3 mL). The resulting solution is stirred atRT and the progress of the reaction is followed by tlc. After reactionoccurs, solvent is removed by evaporation under reduced pressure and theresidue is purified by HPLC, giving the desired compound of Formula I[structure 69, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰and R¹⁴ are H; R¹¹ is 2-(N,N-dimethylamino)ethyl; and R¹² is OAc].

Example 17 See FIG. 16

Preparation of a Formula I compound wherein p is 2, q is 1, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via a 2-aminomethylgroup to the linker, X, and the second ligand, L₂, is thebenzothioazepine moiety linked to X via the amide nitrogen of thethioazepine ring.

Method A

Step 1

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is first added a solution of the benzothiazepine (see FIG. 13structure 41, where R¹² is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol) (1mmol) in dry DMF (3 mL) followed by the linker molecule1-bromomethyl-4-(N-Cbz-N-methyl)aminobenzene (1 mmol) in dry DMF (1 mL).The resulting mixture is stirred and the course of the reaction isfollowed by thin layer chromatography. After reaction occurs, thereaction is quenched with cold dilute aq. Na₂CO₃ and extracted withmethylene chloride. The organic layer is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound is obtained by purification of the crude product by useof HPLC.

Step 2

Ammonium formate (96 mg, 1.5 mmol) and 10% Pd—C (50 mg) are added to asolution of the compound obtained in the preceding reaction in methanol(3 mL) and THF (2 mL). The mixture is stirred at room temperature andthe progress of the reaction is monitored by tlc. After reaction iscomplete, the mixture is filtered through Celite, the filter pad isrinsed with EtOAc, the combined organic layers are washed successivelywith aq. NaHCO₃ and with half-saturated brine, then filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound [(see FIGS. 13 and 16) structure 55, where R¹² is OAc;R¹³ is methyl; and R¹⁴ is H] is obtained by purification of the crudeproduct with the use of HPLC.

Step3

A solution of the compound (structure 55, where R¹² is OAc; R¹³ ismethyl; and R¹⁴ is H) from the preceding reaction (1 mmol) and theN-BOC-1,4-dihydropyridine [structure 25 (Alker, D.; Swanson, A. G.Tetrahedron Lett. 1990, 31, 1479-1482), where PG is t-butyloxycarbonyl(BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (1mmol) and diisopropylethylamine (0.2 mL) in DMF (3 mL) is stirred andwarmed under an inert atmosphere. The progress of the reaction isfollowed by tlc and when reaction is complete, the solution is pouredinto aqueous 5% NaHCO₃ and the aqueous mixture is extracted withmethylene chloride. The organic extract solution is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound is obtained by purification of the crudeproduct by use of HPLC.

Step 4

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 70, where R⁶ andR⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H; R¹² is Oac;and R¹³ is methyl] is obtained by purification of the crude product withthe use of HPLC.

In similar manner, by replacing the 1,4-dihydropyridine of the aboveexample with other ligands of structure 25 and/or the benzothiazepine inthe above example with other ligands of structure 55 and/or by replacing1-bromomethyl-4-(N-Cbz-N-methyl)aminobenzene [in step (1) of thisexample] with other linker molecules, other compounds of Formula I areprepared.

Method B

A solution of the compound (structure 55, where R¹² is OAc; R¹³ ismethyl; and R¹⁴ is H) from the preceding reaction (1 mmol) and the1,4-dihydropyridine [structure 25 (Alker, D.; Swanson, A. G. TetrahedronLett. 1990, 31, 1479-1482), where PG is H; R⁶ and R⁸ are methyl; R⁷ isethyl; R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol) and diisopropylethylamine (0.2mL) in DMF (3 mL) is stirred and warmed under an inert atmosphere. Theprogress of the reaction is followed by tlc and when reaction iscomplete, the solution is poured into aqueous 5% NaHCO₃ and the aqueousmixture is extracted with methylene chloride. The organic extractsolution is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired Formula I compound[structure 70, where R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰and R¹⁴ are H; R¹² is Oac; and R¹³ is methyl] is obtained bypurification of the crude product by use of HPLC.

Example 18 See FIG. 16

Preparation of a Formula I compound wherein p is 2, q is 1, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via a2-hydroxymethyl group to the linker, X, and the second ligand, L₂, isthe benzothioazepine moiety linked to X via the amide nitrogen of thethioazepine ring

Method A

Step 1

A solution, cooled to the temperature of an ice-water bath, containingthe N-BOC-1,4-dihydropyridine (structure 47 [see example 16, method A,step 2)] where PG is BOC; R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl;and R¹⁰ is H} (1 mmol), triphenylphosphine (1.5 mmol), and carbontetrabromide (2 mmol) in CH₂Cl₂ (10 mL) is prepared and is stirred. Thecooling bath is removed and the solution is stirred at room temperature.The progress of the reaction is followed by tlc and after reactionoccurs, the solution is diluted with additional CH₂Cl₂, washed withaqueous 5% NaHCO₃, with water and with half-saturated brine. The organiclayer is separated, dried (Na₂SO₄), filtered and concentrated underreduced pressure to give the crude product. The desired compound(structure 59 where PG is t-butyloxycarbonyl (BOC); R⁶ and R⁸ aremethyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product by use of HPLC.

Step2

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the N-BOC-1,4-dihydropyridine [structure 59,where PG is t-butyloxycarbonyl (BOC); R⁶ and R⁸ are methyl; R⁷ is ethyl;R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol) in DMF (3 mL) followed by a solutionof the benzothiazepine (structure 41, where R¹² is OAc; R¹³ is methyl;and R¹⁴ is H) (1 mmol) in dry DMF (3 mL). The resulting mixture isstirred and the course of the reaction is followed by thin layerchromatography. After reaction occurs, the reaction is quenched withcold dilute aq. Na₂CO₃ and extracted with methylene chloride. Theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound is obtained bypurification of the crude product by use of HPLC.

Step 3

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 71, where R⁶, R⁸and R¹³ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H; R¹² isOAc] is obtained by purification of the crude product with the use ofHPLC.

In similar manner, by replacing the 1,4-dihydropyridine of the aboveexample with other ligands of structure 59 and/or the benzothiazepine inthe above example with other ligands of structure 41 and/or by replacing1-bromomethyl-4-benzyloxybenzene [in example 16] with other linkermolecules, other compounds of Formula I are prepared.

Method B

Step 1

A solution, cooled to the temperature of an ice-water bath, containingthe 1,4-dihydropyridine {structure 47 [see example 16, method B, step 2]where PG is H; R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ isH} (1 mmol), triphenylphosphine (1.5 mmol), and carbon tetrabromide (2mmol) in CH₂Cl₂ (10 mL) is prepared and is stirred. The cooling bath isremoved and the solution is stirred at room temperature. The progress ofthe reaction is followed by tlc and after reaction occurs, the solutionis diluted with additional CH₂Cl₂, washed with aqueous 5% NaHCO₃, withwater and with half-saturated brine. The organic layer is separated,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound (structure 59 where PG is H; R⁶and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained bypurification of the crude product by use of HPLC.

Step 2

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the 1,4-dihydropyridine [structure 59, wherePG is H; R⁶ and R⁸ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; and R¹⁰ is H] (1mmol) in DMF (3 mL) followed by a solution of the benzothiazepine(structure 41, where R¹² is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol)in dry DMF (3 mL). The resulting mixture is stirred and the course ofthe reaction is followed by thin layer chromatography. After reactionoccurs, the reaction is quenched with cold dilute aq. Na₂CO₃ andextracted with methylene chloride. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 71, where R⁶, R⁸,and R¹³ are methyl; R⁷ is ethyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H; Ra¹² isOAc] is obtained by purification of the crude product by use of HPLC.

In similar manner, by replacing the 1,4-dihydropyridine of the aboveexample with other ligands of structure 59 and/or the benzothiazepine inthe above example with other ligands of structure 41 and/or by replacing1-bromomethyl-4-benzyloxybenzene [in example 16, Method A, Part A(1.)]with other linker molecules, other compounds of Formula I are prepared.

Example 19 See FIG. 16

Preparation of a Formula I compound wherein p is 2, q is 1, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via the 3-carboxylgroup to the linker, X, and the second ligand, L₂, is thebenzothioazepine moiety linked toX via the amide nitrogen of thethioazepine ring.

Method A

Step 1

A solution of N-BOC-1,4-dihydropyridine (structure 36, where PG ist-butyloxycarbonyl (BOC); R², R⁶ and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰is H] (1 mmol), the linker molecule 1-hydroxymethyl-4-bromomethylbenzene(1 mmol), and 4-dimethylaminopyridine (10 mg) in CH₂C₂ (5 mL) isprepared under argon in a flask equipped with magnetic stirrer and adrying tube. To this solution is added dicyclohexylcarbodiimide (solid,2.2 mmol). The progress of the reaction is followed by tlc and afterreaction occurs, the reaction solution is quenched in water, aqueoussodium bicarbonate is added and the aqueous mixture is extracted withmethylene chloride. The organic layer is washed with aqueous Na₂CO₃ andwith H₂O, dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound (structure 72,where PG is t-butyloxycarbonyl (BOC); R², R⁶ and R⁸ are methyl; R⁹ is2-Cl; and R¹⁰ is H) is obtained by purification of the crude productwith the use of HPLC.

Step 2

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the benzothiazepine (structure 41, where R¹²is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol) in dry DMF (3 mL) followedby a solution of the N-BOC-1,4-dihydropyridine [structure 72, where PGis t-butyloxycarbonyl (BOC); R², R⁶ and R⁸ are methyl; R⁹ is 2-Cl; andR¹⁰ is H] (1 mmol) in DMF (3 mL). The resulting mixture is stirred andthe course of the reaction is followed by thin layer chromatography.After reaction occurs, the reaction is quenched with cold dilute aq.Na₂CO₃ and extracted with methylene chloride. The organic layer is dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound is obtained by purification of thecrude product by use of HPLC.

Step 3

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 73, where R², R⁶,R⁸ and R¹³ are methyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H; and R¹² is OAc] isobtained by purification of the crude product with the use of HPLC.

In similar manner, by replacing the 1,4-dihydropyridine of the aboveexample with other ligands of structure 72 and/or the benzothiazepine inthe above example with other ligands of structure 41 and/or by replacing1-hydroxymethyl-4-bromomethylbenzene with other linker molecules, othercompounds of Formula I are prepared.

Method B

Step 1

A solution of 1,4-dihydropyridine (structure 36, where PG is H; R², R⁶and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol), the linkermolecule 1-hydroxymethyl-4-bromomethylbenzene (1 mmol), and4-dimethylaminopyridine (10 mg) in CH₂Cl₂ (5 mL) is prepared under argonin a flask equipped with magnetic stirrer and a drying tube. To thissolution is added dicyclohexylcarbodiimide (solid, 2.2 mmol). Theprogress of the reaction is followed by tlc and after reaction occurs,the reaction solution is quenched in water, aqueous sodium bicarbonateis added and the aqueous mixture is extracted with methylene chloride.The organic layer is washed with aqueous Na₂CO₃ and with H₂O, dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound (structure 72, where PG is H; R², R⁶and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H) is obtained by purificationof the crude product with the use of HPLC.

Step 2

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the benzothiazepine (structure 41, where R¹²is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol) in dry DMF (3 mL) followedby a solution of the dihydropyridine [structure 72, where PG is H; R²,R⁶ and R⁸ are methyl; R⁹ is 2-Cl; and R¹⁰ is H] (1 mmol) in DMF (3 mL).The resulting mixture is stirred and the course of the reaction isfollowed by thin layer chromatography. After reaction occurs, thereaction is quenched with cold dilute aq. Na₂CO₃ and extracted withmethylene chloride. The organic layer is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired Formula I compound [structure 73, where R², R⁶, R⁸ and R¹³ aremethyl; R⁹ is 2-Cl; R¹⁰ and R¹⁴ are H; and R¹² is OAc] is obtained bypurification of the crude product by use of HPLC.

In similar manner, by replacing the 1,4-dihydropyridine of the aboveexample with other ligands of structure 72 and/or the benzothiazepine inthe above example with other ligands of structure 41 and/or by replacing1-hydroxymethyl-4-bromomethylbenzene with other linker molecules, othercompounds of Formula I are prepared.

Example 20 See FIG. 17

Preparation of a Formula I compound wherein p is 2, q is 7, and oneligand, L₁, is the 1,4-dihydropyridine moiety linked via a 6-amino groupto the linker, X, and the second ligand, L₂, is the benzthioazepinemoiety linked to X via the hydroxyl function of the phenolic ring.

Method A

Step 1

A solution, cooled to the temperature of an ice-water bath, containingthe benzothioazepine {structure 52 [see example 9], where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H} (1 mmol),triphenylphosphine (1.5 mmol), and carbon tetrabromide (2 mmol) inCH₂Cl₂ (10 mL) is prepared and is stirred. The cooling bath is removedand the solution is stirred at room temperature. The progress of thereaction is followed by tlc and after reaction occurs, the solution isdiluted with additional CH₂Cl₂, washed with aqueous 5% NaHCO₃, withwater and with half-saturated brine. The organic layer is separated,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound [structure 66 where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] is obtained bypurification of the crude product by use of HPLC.

Step 2

A solution of the compound [structure 66 where R¹¹ is2-(N,N-dimethylamino)-ethyl; R¹² is OAc; and R¹⁴ is H] (1 mmol) preparedin the preceding reaction, the N-BOC-1,4-dihydropyridine [structure 75,where PG is t-butyloxycarbonyl (BOC)] (1 mmol), anddiisopropylethylamine (0.2 mL) in DMF (5 mL) is stirred and warmed underan inert atmosphere. The progress of the reaction is followed by tlc andwhen reaction is complete, the solution is poured into aqueous 5% NaHCO₃and the aqueous mixture is extracted with methylene chloride. Theorganic extract solution is dried (Na₂SO₄), filtered and concentratedunder reduced pressure to give the crude product. The desired compoundis obtained by purification of the crude product by use of HPLC.

Step 3

A solution of the product from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound of Formula I [structure 76, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] is obtained bypurification of the crude product with the use of HPLC.

In similar manner, by replacing the benzothiazepine in the above examplewith other ligands of structure 66 and/or by replacing the linkermolecule used to prepare 52 with other linker molecules, other compoundsof Formula I are prepared.

Method B

Step 1

A solution, cooled to the temperature of an ice-water bath, containingthe benzothioazepine {structure 52 [see example 9, method A, step 2],where R¹¹ is 2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H} (1mmol), triphenylphosphine (1.5 mmol), and carbon tetrabromide (2 mmol)in CH₂Cl₂ (10 mL) is prepared and is stirred. The cooling bath isremoved and the solution is stirred at room temperature. The progress ofthe reaction is followed by tlc and after reaction occurs, the solutionis diluted with additional CH₂Cl₂, washed with aqueous 5% NaHCO₃, withwater and with half-saturated brine. The organic layer is separated,dried (Na₂SO₄), filtered and concentrated under reduced pressure to givethe crude product. The desired compound [structure 66 where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] is obtained bypurification of the crude product by use of HPLC.

Step 2

A solution of the compound [structure 66 where R¹⁰ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H] (1 mmol) preparedin the preceding reaction, the dihydropyridine [structure 75, where PGis H] (1 mmol), and diisopropylethylamine (0.2 mL) in DMF (5 mL) isstirred and warmed under an inert atmosphere. The progress of thereaction is followed by tlc and when reaction is complete, the solutionis poured into aqueous 5% NaHCO₃ and the aqueous mixture is extractedwith methylene chloride. The organic extract solution is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired Formula I compound [structure 76, where R¹¹ is2-(N,N-dimethylamino)ethyl; R¹² is OAc; and R¹⁴ is H]compound isobtained by purification of the crude product by use of HPLC.

In similar manner, by replacing the benzothiazepine in the above examplewith other ligands of structure 66 and/or by replacing the linkermolecule used to prepare 52 with other linker molecules, other compoundsof Formula I are prepared.

Example 21 See FIG. 18

Preparation of a compound in which ligand, L₁, is linked directly toligand, L₂, where L₁ is the amlodipine moiety and L₂ is the diltiazemmoiety and where R=H.

Method A

Step 1

A mixture of NaH (1.1 mmol) and DMF (1 mL) is prepared under an inertatmosphere in a flask equipped with a stirring bar and a drying tube. Tothis is added a solution of the benzothiazepine (structure 41 where R¹²is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol) in dry DMF (3 mL) followedby 1,2-dibromoethane (10 mmol). The resulting mixture is stirred and thecourse of the reaction is followed by thin layer chromatography. Afterreaction occurs, the reaction is quenched with cold dilute aq. Na₂CO₃and extracted with methylene chloride. The organic layer is dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound (structure 88) is obtained bypurification of the crude product by use of HPLC.

Step 2

A solution of the compound (structure 88) (1 mmol) prepared in thepreceding reaction, N-BOC-amlodipine [structure 87, where PG ist-butyloxycarbonyl (BOC)] (1 mmol), and diisopropylethylamine (0.2 mL)in DMF (5 mL) is stirred and warmed under an inert atmosphere. Theprogress of the reaction is followed by tlc and when reaction iscomplete, the solution is poured into aqueous 5% NaHCO₃ and the aqueousmixture is extracted with methylene chloride. The organic extractsolution is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound (structure 89)is obtained by purification of the crude product by use of HPLC.

Step 3

A solution of the product (structure 89) from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound (structure 90) is obtained by purificationof the crude product with the use of HPLC.

Method B

A solution of the compound (structure 88) (1 mmol) prepared in thepreceding reaction, amlodipine (structure 87, where PG is H) (1 mmol),and diisopropylethylamine (0.2 mL) in DMF (5 mL) is stirred and warmedunder an inert atmosphere. The progress of the reaction is followed bytlc and when reaction is complete, the solution is poured into aqueous5% NaHCO₃ and the aqueous mixture is extracted with methylene chloride.The organic extract solution is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound (structure 90) is obtained by purification of the crudeproduct by use of HPLC.

Example 22 See FIG. 19

Preparation of a compound in which ligand, L₁, is linked directly toligand, L₂, where L¹ is the amlodipine moiety and L₂ is the diltiazemmoiety

Method A

Step 1

A solution of compound (structure 89) (1 mmol) and paraformaldehyde (2mmols) in methanol (4 mL) is stirred and is acidified with acetic acidto pH 6.6 (pH meter) under a nitrogen atmosphere. Sodiumcyanoborohydride (1.1 mmol) is then added and stirring is continued. Thecourse of the reaction is followed by thin layer chromatography. Afterreaction occurs, the reaction solution is quenched in water and the pHof the aqueous mixture is adjusted to greater than 10 with aqueous NaOH.The mixture is extracted with ether, the organic extracts are washedwith half-saturated saline, dried (Na₂SO₄), filtered and concentratedunder reduced pressure to give the crude product. The desired compound(structure 91) is obtained by purification of the crude product by useof HPLC.

Step 2

A solution of the product (structure 91) from the preceding reaction andtrifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred at roomtemperature. The progress of the reaction is followed by tlc. Afterreaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired compound (structure 92) is obtained by purificationof the crude product with the use of HPLC.

Method B

Step 1

A solution of the compound (structure 89) (1 mmol) and paraformaldehyde(5 mmol) in ethanol (5 mL) is stirred with 10% Pd—C (20 mg) under ahydrogen atmosphere. The progress of the reaction is followed by tlc.After reaction occurs, the mixture is filtered through Celite, thefilter pad is washed with ethanol, and the filtrates are concentratedunder reduced pressure. The desired compound (structure 91) is obtainedby purification of the crude product with the use of HPLC and isconverted to structure 92 as described above in Method A.

(Method C)

A solution of the compound (structure 89) (1 mmol) in ether (5 mL) isadded slowly to a vigorously stirred mixture of a solution methyl iodide(1 mmol) in ether (5 mL) and a solution of Na₂CO₃ in H₂O (1 mL). Theprogress of the reaction is followed by tlc. After reaction is complete,the mixture is washed with additional aqueous Na₂CO₃ and with H₂O, theorganic layer is dried (Na₂SO₄), filtered and concentrated under reducedpressure to give the crude product. The desired compound (structure 91)is obtained by purification of the crude product with the use of HPLC.The compound (structure 91) is converted to structure 92 as describedabove in Method A.

(Method D)

A solution of the compound (structure 90) (1 mmol) and paraformaldehyde(5 mmol) in ethanol (5 mL) is stirred with 10% Pd—C (20 mg) under ahydrogen atmosphere. The progress of the reaction is followed by tlc.After reaction occurs, the mixture is filtered through Celite, thefilter pad is washed with ethanol, and the filtrates are concentratedunder reduced pressure. The desired compound (structure 92) is obtainedby purification of the crude product with the use of HPLC.

Example 23 See FIG. 20

Preparation of a compound in which ligand, L₁, is linked directly toligand, L₂, where L¹ is the amlodipine moiety and L₂ is the diltiazemmoiety and where R=H.

Step 1

A mixture of ethanolamine (0.1 mol), di-tert-butylcarbonate (0.15 mol),dioxane (50 mL) and aq. 2 N NaOH (25 mL) is stirred at RT for 24 hr. Thedioxane is removed by evaporation under reduced pressure. Water (50 mL)is added to the aqueous mixture and the mixture is extracted with CH₂Cl₂(4×25 mL). The combined organic layers are dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. PureN-BOC-ethanolamine is obtained by purification of the crude product withthe use of flash chromatography over silica gel.

Step 2

t-Butyldimethylsilyl chloride(0.4 mol) is added to a solution ofN-BOC-ethanolamine (0.1 mole) and imidazole (0.1 mol) in dry pyridine(75 mL) and the resulting solution is stirred at RT. The progress of thereaction is followed by tlc. When reaction is complete, water (5 mL) isadded to the solution which is then concentrated by evaporation underreduced pressure (>25 mm Hg, 30° C.). The residue is dissolved in EtOAcand the solution is extracted with saturated aq. CuSO₄ to removeresidual pyridine. The EtOAc solution is washed with water, dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The pure product, N-BOC-ethanolamine-O-TBDMS, is obtainedby purification of the crude product by flash chromatography over silicagel.

Step 3

A solution of N-BOC-ethanolamine-O-TBDMS (0.05 mol) in dry DMF (3 mL) isadded dropwise to a stirred mixture of NaH (0.05 mol) and dry DMF (10mL) under an inert atmosphere. The resulting mixture is stirred for 1 hrand then is tranferred by cannulation to a stirred solution of1,2-dibromoethane (0.3 mol) in dry DMF (10 mL). The resulting solutionis stirred and the progress of the reaction is followed by tlc. Afterreaction occurs, the reaction solution is quenched with aqueous 5%NaHCO₃ (100 mL) and brine (100 mL). The mixture is extracted with CH₂Cl₂(4×25 mL) and the combined organic extracts are back-washed with water(3×). The organic layer is dried (Na₂SO₄), filtered and concentratedunder reduced pressure to give the crude product. PureN-BOC-N-(2-bromoethyl) ethanolamine-O-TBDMS is obtained by purificationof the crude product with the use of flash chromatography over silicagel.

Step 4

A mixture of NaH (1.1 mmol) and dry DMF (1 mL) is prepared under aninert atmosphere in a flask equipped with a stirring bar and a dryingtube. To this is added a solution of the benzothiazepine (structure 41where R¹² is OAc; R¹³ is methyl; and R¹⁴ is H) (1 mmol) in dry DMF (3mL). Then a solution of N-BOC-N-(2-bromoethyl)ethanolamine-O-TBDMS (1mmol) in dry DMF (2 mL) is added and the resulting mixture is stirredand monitored for reaction by tlc. After reaction occurs, the reactionsolution is quenched with aqueous 5% NaHCO₃ (25 mL) and brine (25 mL).The mixture is extracted with CH₂Cl₂ (4×20 mL) and the combined organicextracts are back-washed with water (3×). The organic layer is dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound (structure 93) is obtained bypurification of the crude product with the use of HPLC.

Step 5

A solution of the product (structure 93) (1 mmol) from the precedingreaction and Et₃N-(HF)₃ in MeCN (5 mL) is stirred at room temperature.After reaction occurs as detected by tlc, the solution is diluted withEtOAc and then washed with water-brine. The organic layer is dried(Na₂SO₄), filtered and concentrated under reduced pressure to give thecrude product. The desired compound (structure 94) is obtained bypurification of the crude product with the use of HPLC.

Step 6

A mixture of NaH (2.1 mmol) and dry DMF (1 mL) is prepared under aninert atmosphere in a flask equipped with a stirring bar and a dryingtube. To this is added a solution of the compound (structure 94)prepared in the preceding reaction (1 mmol) in dry DMF (3 mL). Then asolution of the 1,4-dihydropyridine 26 (where PG=H) (1 mmol) in dry DMF(2 mL) is added and the resulting mixture is stirred and monitored forreaction by tlc. After reaction occurs, the reaction solution isquenched water (25 mL) and brine (25 mL). The mixture is extracted withCH₂Cl₂ (4×20 mL) and the combined organic extracts are back-washed withwater (3×). The organic layer is dried (Na₂SO₄), filtered andconcentrated under reduced pressure to give the crude product. Thedesired compound (structure 89, where PG=H; R=BOC) is obtained bypurification of the crude product with the use of HPLC.

Step 7

A solution of the product (structure 89, PG=H; R=BOC) from the precedingreaction and trifluoroacetic acid (3 mL) in CH₂Cl₂ (5 mL) is stirred atroom temperature. The progress of the reaction is followed by tlc. After(D reaction occurs, more CH₂Cl₂ is added and the solution is washed withaqueous Na₂CO₃ and with H₂O. The organic layer is dried (Na₂SO₄),filtered and concentrated under reduced pressure to give the crudeproduct. The desired Formula I compound (structure 90) is obtained bypurification of the crude product with the use of HPLC.

While the present invention has been described with reference to thespecific embodiments thereof, it should be understood by those skilledin the art that various changes may be made and equivalents may besubstituted without departing from the true spirit and scope of theinvention. In addition, many modifications may be made to adapt aparticular situation, material, composition of matter, process, processstep or steps, to the objective spirit and scope of the presentinvention. All such modifications are intended to be within the scope ofthe claims appended hereto.

All of the publications, patent applications and patents cited in thisapplication are herein incorporated by reference in their entirety tothe same extent as if each individual publication, patent application orpatent was specifically and individually indicated to be incorporated byreference in its entirety.

1. A compound of the formula:L-X-L or a pharmaceutically-acceptable salt thereof; wherein one L is aligand of formula (vi):

and the other L is a ligand independently selected from the groupconsisting of formula (i), (ii), (iii), (iv), (v) and (vi):

and X is a linker of the formula:—X′-Z-(Y′-Z)_(m)-Y″-Z-X′— in which: m is an integer from 0 to 20; X′ ateach separate occurrence is —O—, —S—, —S(O)—, —S(O)2-, —NR—, —NRR′,—C(O)—, —C(O)O—, —C(O)NH—, —C(S)—, —C(S)O—, —C(S)NH— or a covalent bond,where R and R′ at each separate occurrence are as defined below for R′and R″; Z is at each separate occurrence selected from alkylene,substituted alkylene, alkylalkoxy, cycloalkylene, substitutedcycloalkylene, alkenylene, substituted alkenylene, alkynylene,substituted alkynylene, cycloalkenylene, substituted cycloalkenylene,arylene, substituted arylene, heteroarylene, heterocyclene, substitutedheterocyclene, crown compounds or a covalent bond; Y′ and Y″ at eachseparate occurrence are selected from —S—S—, a covalent bond or astructure selected from the following group:

in which: n is 0, 1 or 2; and R′ and R″ at each separate occurrence areselected from hydrogen, alkyl, substituted alkyl, cycloalkyl,substituted cycloalkyl, alkenyl, substituted alkenyl, alkynyl,substituted alkynyl, aryl, heteroaryl or heterocyclic.
 2. The compoundaccording to claim 1, wherein the compound is of the followingstructure:


3. A pharmaceutical composition comprising a pharmaceutically acceptableexcipient and a therapeutically effective amount of a compound of claim1 or 2.